Lyme disease is caused by the bacterium Borrelia burgdorferi. While it is still the most common vector-borne disease in the country, it is also transmitted in other ways. Infection occurs through blood transfusion, organ transplant, direct contact and by congenital means.

Hematogenous dissemination of organisms occurs in many spirochetal diseases, including Lyme disease and syphilis. Although syphilis has been transmitted by transfusion, in the vast majority of cases, only fresh blood products were involved, in part because Treponema pallidum survives poorly when refrigerated in citrated blood. Because of the rising incidence of Lyme disease in certain areas, whether its causative agent, Borrelia burgdorferi, could survive under blood banking conditions was studied. Dilutions of stock cultures of two strains of B. burgdorferi were inoculated into samples of citrated red cells (RBCs). Viable spirochetes were recovered from RBCs inoculated with 10(6) organisms per mL, after refrigeration for as long as 6 weeks. It is concluded that B. burgdorferi may survive storage under blood banking conditions and that transfusion-related Lyme disease is theoretically possible.

In order to determine if Peromyscus spp. could become infected with the Lyme disease spirochete (Borrelia burgdorferi) by direct inoculation and to determine the duration of spirochetemia, 4 P. leucopus and 5 P. maniculatus were inoculated by the intramuscular, intraperitoneal, and subcutaneous routes with an isolate of B. burgdorferi obtained from the blood of a trapped wild P. leucopus from Camp McCoy, Wisconsin. All of the mice developed antibodies to B. burgdorferi which reached a peak indirect immunofluorescent (IFA) geometric mean antibody titer of 10 log2 21 days post-inoculation. B burgdorferi was recovered from the blood of 1 P. maniculatus 21 days post-inoculation. One uninfected Peromyscus of each species was housed in the same cage with the infected Peromyscus as a contact control. Both of the contact controls developed IFA B. burgdorferi antibodies by day 14, indicating contact infection. To determine if B. burgdorferi was being transmitted by direct contact, 5 uninfected P. leucopus and 5 uninfected P. maniculatus were caged with 3 B. burgdorferi infected P. leucopus and 3 infected P. maniculatus, respectively. Each of these contact-exposed P. leucopus and P. maniculatus developed antibodies to B. burgdorferi, and B. burgdorferi was isolated from the blood of 1 contact-exposed P. maniculatus 42 days post-initial contact. These findings show that B. burgdorferi can be transmitted by direct contact without an arthropod vector.

Abstract
Babesiosis was reported in a California resident who received a transfusion of blood products collected in the disease-endemic northeastern region of the United States. Babesiosis should be considered year-round in the diagnosis of febrile and afebrile patients with abnormal blood cell counts who have received blood products from disease-endemic areas.

The aim of the study was to determine the level of infection in mosquitoes with spirochetes Borrelia burgdorferi sensu lato in the woody areas of Szczecin. The mosquitoes were collected from May to September 2003. The spirochetes, Borrelia burgdorferi s. l., present in mosquitoes were detected in mosquitoes with indirect immunofluorescence assay (IFA) using rabbit anti-Borrelia burgdorferi antibodies and goat anti-rabbit IgG marked with fluorescein isocyanate (FITC). A total of 1557 females and 58 males were collected. They represented the genera Aedes (63%) and Culex (37%). The infection level of the mosquitoes from the area studied amounted to 1.7%. The results of the present study confirm the potential of these arthropods to spread Lyme borreliosis.

Lyme disease, being a spirochete with pathology similar to syphilis, is often found difficult to treat due to the spirochete invading sanctuary sites and displaying pleomorphic characteristics such as a cyst (L-form). Because a significant portion of sexually active couples present to my office with Lyme disease, with only one partner having a history of tick exposure, the question of possible secondary (sexual)vector of transmission for the spirochete warrents inquiry. Additionally, sexually active couples seem to have a marked propensity for antibiotic failure raising the question of sexually active couples re-infecting themselves through intimate contact.

METHODS:

Lyme spirochetes/DNA have been recovered from stored animal semen. Recovery of spirochete DNA from nursing mother's breast milk and umbilical cord blood by PCR (confirmed by culture/microscopy), have been found in samples provided to my office.

RESULTS:

Surprisingly, initial laboratory testing of semen samples provided by male Lyme patients (positive by western blot/PCR in blood) and the male sexual partner of a Lyme infected female patient were positive approximately 40% of the time. PCR recovery of Lyme DNA nucleotide sequences with microscopic confirmation of semen samples yielded positive results in 14/32 Lyme patients (13 male semen samples and 1 vaginal pap). ALL positive semen/vaginal samples in patients with known sexual partners resulted in
positive Lyme titres/PCR in their sexual partners. 3/4 positive semen patients had no or unknown sexual partners to be tested.

These preliminary findings warrant further study. Current a statistical design study to evaluate the possibility of sexual transition of the spirochete is being undertaken. Our laboratory studies confirm the existence of Lyme spirochetes in semen/vaginal secretions. Whether or not further clinical studies with a larger statistical group will support the hypothesis of sexual transmission remains to be seen. A retrospective clinical study is also underway. We are reviewing the medical records, collecting semen samples of patients who were
previously diagnosed with current and previously treated Lyme disease are being asked to provide semen,pap and blood samples for extensive laboratory testing.

CONCLUSION;

With the initially impressive data, we feel the subsequent statistical study on the sexual transmission of the Lyme spirochete will illuminate a much broader spectrum of public health concerns associated with the disease than the originally accepted tick borne vector.

Infectious Diseases Section and Department of Transfusion Medicine, National Institutes of Health, Bethesda, MD.
Following the devastating effects of blood-transmitted human immunodeficiency virus (HIV), blood establishments have become increasingly vigilant for the emergence or re-emergence of new threats to the safety of the blood supply. Many agents have fulfilled the broad definition of emerging blood-transmitted infections, including West Nile virus (WNV), Trypanosoma cruzi, Plasmodium spp., Babesia spp., parvovirus B19, dengue virus, and the prions that cause variant Creutzfeld-Jacob disease (vCJD). Other agents such as human herpes virus- 8 (HHV-8-Kaposi's sarcoma virus) and Borellia (Lyme disease) and, perhaps, avian flu virus, are known to have a viremic phase, but have not yet been proved to be transfusion-transmitted. In the wake of these threats, transfusion services use a variety of donor screening interventions, including serologic assays, nucleic acid assays, and geographic exclusions based on potential exposure. The ultimate safeguard may be a pre-emptive pathogen inactivation strategy that will disrupt all nucleic acid-containing agents (though not prions).

Considerable effort and resources have been invested in this arena, but currently no single technique is effective for inactivation of both liquid and cellular blood products and toxicity issues have not been completely resolved. The blood supply is remarkably safe with the risk of major pathogens such as hepatitis C virus (HCV) and HIV now reduced to less than one transmission per 2 to 3 million exposures. However, to approach near-zero infectious disease risk for emerging and re-emerging pathogens, new strategies such as pathogen inactivation or multi-pathogen microarray technology will need to be developed or refined.

RECOVERY OF LYME SPIROCHETES BY
PCR IN SEMEN SAMPLES OF PREVIOUSLY
DIAGNOSED LYME DISEASE PATIENTS
Presented by Dr. Gregory Bach, at the International
Scientific Conference on Lyme Disease, April, 2001.

OBJECTIVE:
Lyme disease, being a spirochete with pathology similar to syphilis, is often found difficult to treat due to the spirochete invading sanctuary sites and displaying pleomorphic characteristics such as a cyst (L-form). Because a significant portion of sexually active couples present to my office with Lyme disease, with only one partner having a history of tick exposure, the question of possible secondary (sexual)vector of transmission for the spirochete warrents inquiriy.
Additionally, sexually active couples seem to have a marked propensity for antibiotic failure raising the question of sexually active couples re-infecting themselves through intimate contact.
METHODS:
Lyme spirochetes/DNA have been recovered from stored animal semen. Recovery of spirochete DNA from nursing mother's breast milk and unbilical cord blood by PCR (confirmed by culture/microscopy), have been found in samples provided to my office.

ALL positive semen/vaginal samples in patients with known sexual partners resulted in positive Lyme titers/PCR in their sexual partners. 3/4 positive semen patients had no or unknown sexual partners to be tested. These preliminary findings warrent futher study. Current a statistical design study to evaluate the possibility of sexual transition of the spirochete is being undertaken.

Our laboratory studies confirm the existence of Lyme spirochetes in semen/vaginal secretions. Whether or not further clinical studies with a larger statistical group will support the hypothesis of sexual transmission remains to be seen. A retrospective clinical study is also underway.
We are reviewing the medical records, collecting semen samples of patients who were previously diagnosed with current and previously treated Lyme disease are bing asked to provide semen,pap and blood samples for extensive laboratory testing.

CONCLUSION:
With the initially impressive data, we feel the subsequent statistical sudy on the sexual transmission of the Lyme spirochete will illuminate a much broader sectrum of public health concerns associated with the disease than the originally accepted tick borne vector.

Between June 2002 and July 2003, 987 fleas, representing four species, and 1019 ticks, representing one argasid and eight ixodid species, were collected from Egyptian animals. These arthropods were tested for rickettsial agents using polymerase chain reaction. DNAs from Anaplasma and Ehrlichia spp. were detected in 13 ticks. Previously undescribed Bartonella spp. were detected in 21 fleas. Coxiella burnetii was detected in two fleas and 20 ticks. Rickettsia typhi was detected in 27 fleas from 10 cities. Spotted fever group rickettsiae were detected in both fleas and ticks and included Rickettsia aeschlimanii and an unnamed Rickettsia sp.

A total of 662 samples (winter period: 469; summer period: 193 specimens) of female mosquitoes of the genus Culex, Aedes and Anopheles were collected during the period March 2000-April 2001 from the locality of Vysoke Myto (Eastern Bohemia, Czech Republic). They were examined by dark field microscopy for the presence of spirochetes. The motile spirochetes were observed in 4.2 % of all species of investigated mosquitoes. One spirochetal strain out of the 8 isolation attempts (BRZ14) was obtained (cultivation rate was 12.5 %) and the spirochetal strain was then successfully cultivated and identified using PCR for the presence of Borrelia burgdorferi s.l., and subsequently with the RFLP as genomospecies Borrelia afzelii. This strain was derived from overwintering Culex (Culex) pipiens biotype molestus female mosquitoe. This is apparently one of the sporadic cases of the occurrence of pathogenic borreliae in haematophagous arthropods, other than Ixodes ricinus complex ticks.

The results of the present study indicate that ticks have the highest potential for infecting humans with Lyme disease. Within the area studied, 17.1% of ticks were infected with Borrelia burgdorferi spirochetes. Other authors surveying other regions of Poland revealed lower values of the tick infection. Wegner et al., using also IFA, revealed tick infection of 11.5% in the former Olsztyn province in 1993 [31], and 8.8% at recreational areas of burgdorferi. Bukowska et al. [2] detected the presence of B. burgdorferi in 11.6% of ticks collected in the area of Szczecin in 2000 and 9.6% in 2001, while Michalik et al., collected 16.2% in 1998?1999 in popular recreational.

Other authors, employing PCR method, also obtained lower infection values of ticks. et al. [25] revealed 13% infection of ticks captured in the proximity study by Wodecka [33] on the occurrence of B. burgdorferi in the population of I. ricinus in northwestern Poland in 1998-2001 revealed that infected ticks constituted 9.4%. Skotarczak and Wodecka [26], surveying areas of the Zachodniopomorskie Province in 1996, found that 12% of Ixodes ricinus ticks were infected. Skotarczak [27] detected the presence of the spirochetes in 8.6% of tick specimens collected from the same areas in 1997. In 2000 and 2001, ticks from the Mazury lakes amounting to 6.2 and 2.6%, respectively. On the other hand, values of infection higher than in the present study, were found only in Wielkopolska by Jenek 1995 (24.5%), by Nowosad et al. [20] in 1997?1998 (22.6%). Even though the principal vector of Lyme borreliosis are ticks, the role of hematophagous insects in the epidemiology of this disease cannot be underestimated.

Table 1. Prevalence of infection of ticks Ixodes ricinus and Aëdes with spirochetes Borrelia burgdorferi within the area studied in 2000 and 2001.

Number of specimens collected infected

Prevalence

(%)

Ticks 215 38 17.7

? Nymphs 193 30 15.5

? Adults 22 8 36.4

Aëdes 947 8 0.8

Prevalence of Borrelia burgdorferi in arthropod vectors within Szczecin area 107

Not all borreliosis-affected patients admitted, in their medical interviews, to having contact with ticks [21]. There have been few documented cases of Lyme borreliosis related to insects in Canada [3] and in the USA (Connecticut) [15]. In Sweden, on the other hand, there was a case of erythema migrans observed after a mosquito bite [6]. In Poland there are about 40 species of mosquitoes representing 5 genera: Anopheles, Aëdes, Culex, Culiseta, and Mansonia [12]. Thirty species of those insects have been reported in Szczecin. Of this number 10 species, including those representing the genus Aëdes, are known to transmit microorganisms pathogenic to humans [29]. In the present study, the collected mosquitoes, lured to a human body, were females representing the genus Aëdes, which is consistent with the observations of Lachmajer et al. [12], who noticed that those insects attack in shaded areas, among trees and bushes.

Mosquitoes are annoying insects, especially when they occur in mass numbers. Mosquitoes also transmit various microorganisms pathogenic to humans, among them Borrelia burgdorferi was recovered from mosquitoes representing the genera Aëdes, Culex, and Anopheles [1, 4, 5, 7, 10, 11, 15, 16, 17, 18]. Studies on the occurrence of B. burgdorferi in mosquitoes have been carried out in a number of research centres in the world. Also in Poland, Kubica-Biernat et al. [11] detected B. burgdorferi a 0.5% infection rate. In our earlier study [10], carried out at recreational areas of Szczecin, we detected prevalence values between 0.6? 3.2% in mosquitoes of the genus Aëdes.

It can be concluded from the results of the other authors that the infection frequency of the Lyme borreliosis among humans is related to the percentage of infected ticks or mosquitoes. In the areas of particularly high incidence of human borreliosis (e.g. Connecticut), Borrelia burgdorferi was found with the aid of IFA in 36.2% of ticks, Ixodes scapularis and in 9.5?11.1% of mosquitoes of the genus Aëdes [16]. On the other hand, in Moravia, Hubalek et al. [7] observed 20.4% of Ixodes ricinus and 4.1% of mosquitoes infected with Borrelia burgdorferi.

The low percentage of infected mosquitoes compared to infected ticks may be related to the 2-week survival period of the spirochetes in the organism of those insects [18]. There has been no report on experimentally proven cases of transovarian or transstadial transmission of the spirochetes,although detection of B. burgdorferi in mosquito larvae by Zakovska [34] may indicate their transovarian route.

Borrelia burgdorferi was identified as the etiological agent of Lyme disease in 1982. This Gram-negative spirochete is classified in the order Spirochaetales and the family Spirochaetaceae. The pathogen is fastidious, microaerophilic, mesophilic and metabolises glucose through the Embden-Meyerhof pathway. A generation time of 11 to 12 h at 37 degrees C in Barbour-Stoenner-Kelly medium has been reported. Lyme disease, named after Lyme in Connecticut, is distributed globally. It is the most commonly reported vector-borne disease in the United States, where the incidence is highest in the eastern and midwestern states. Since establishment of national surveillance in 1982, there has been a nine-fold increase in the number of cases reported to the U.S. Centers for Disease Control. The deer tick of the genus Ixodes is the primary vector of Lyme borreliosis. The tick may become infected with B. burgdorferi, by feeding on an infected host, at any point in its 2-year life cycle which involves larval, nymphal and adult stages. The infection rate in deer ticks may be as high as 40% in endemic areas. The primary vertebrate reservoirs for Ixodes are the white-footed mouse (Peromyscus leucopus) and the white-tailed deer (Odocopileus virginianus). Dairy cattle and other food animals can be infected with B. burgdorferi and hence some raw foods of animal origin might be contaminated with the pathogen. Recent findings indicate that the pathogen may be transmitted orally to laboratory animals, without an arthropod vector. Thus, the possibility exists that Lyme disease can be a food infection. In humans, the symptoms of Lyme disease, which manifest themselves days to years after the onset of infection, may involve the skin, cardiac, nervous and/or muscular systems, and so misdiagnosis can occur.

In order to determine if Peromyscus spp. could become infected with the Lyme disease spirochete (Borrelia burgdorferi) by direct inoculation and to determine the duration of spirochetemia, 4 P. leucopus and 5 P. maniculatus were inoculated by the intramuscular, intraperitoneal, and subcutaneous routes with an isolate of B. burgdorferi obtained from the blood of a trapped wild P. leucopus from Camp McCoy, Wisconsin.

All of the mice developed antibodies to B. burgdorferi which reached a peak indirect immunofluorescent (IFA) geometric mean antibody titer of 10 log2 21 days post-inoculation. B burgdorferi was recovered from the blood of 1 P. maniculatus 21 days post-inoculation.

One uninfected Peromyscus of each species was housed in the same cage with the infected Peromyscus as a contact control. Both of the contact controls developed IFA B. burgdorferi antibodies by day 14, indicating contact infection.

Each of these contact-exposed P. leucopus and P. maniculatus developed antibodies to B. burgdorferi, and B. burgdorferi was isolated from the blood of 1 contact-exposed P. maniculatus 42 days post-initial contact.

These findings show that B. burgdorferi can be transmitted by direct contact without an arthropod vector.

Ludwig Boltzmann Institute for Dermato-Venerological Serodiagnosis, University of Vienna, Austria.

Current laboratory diagnosis of Lyme borreliosis relies on tests for the detection of antibodies to Borrelia burgdorferi with known limitations. By using a simple extraction procedure for urine samples, B. burgdorferi DNA was amplified by a nested PCR with primers that target the specific part of the flagellin gene. To control possible inhibition of the enzyme (polymerase), a special assay using the same primers was developed. We examined 403 urine samples from 185 patients with skin manifestations of Lyme borreliosis. Before treatment, B. burgdorferi DNA was detected in 88 of 97 patients with Lyme borreliosis. After treatment, all but seven patients became nonreactive. Six of these seven persons suffered from intermittent migratory arthralgias or myalgias, and one from acrodermatitis chronica atrophicans. Two of 49 control patients with various dermatologic disorders and none out of 22 presumably healthy persons were reactive in the PCR. In addition to urine, breast milk from two lactating women with erythema migrans was tested and also found reactive. Borrelia burgdorferi DNA can be detected with high sensitivity (91%) by a nested PCR in urine of patients with Lyme borreliosis. In addition, this test can be a reliable marker for the efficacy of treatment.

Recent research by Penn biology professor Dustin Brisson suggests that chipmunks and two shrew species account for nearly three-quarters of carriers of ticks infected with Lyme disease.

The widely held belief was that mice were the main animal carriers of the disease.

The research was conducted in Hudson Valley with Daniel Dykhuizen of Stony Brook University and Richard Ostfeld of the Institute of Ecosystem Studies.

In a University press release Brisson said, "The majority of zoonotic diseases, those that can be transmitted from wild or domestic animals to humans, are generally assumed to have one natural animal host."

Though deer are often associated with transmitting Lyme-disease infected ticks to humans, the insects are rarely infected with the bacteria from the deer's blood.

Rather, ticks harbor the disease after they first drink the blood of a vertebrate, which Brisson's research shows is often from chipmunks and shrews in addition to mice.

Mice were originally thought to be the primary carriers because nearly 90 percent of ticks feeding on an infected mouse contract the disease.

In 2004, a total of 618 Ixodes ricinus ticks fed on humans were collected by physicians throughout Thuringia. The prevalence rates of Borrelia burgdorferi sensu lato (s.l.) genotypes were determined by nested PCR and restriction fragment length polymorphism, targeting a 0.8-kb fragment of the ospA gene. The total prevalence of B. burgdorferi s.l. was 6.1%. B. afzelii was found in 67%, B. valaisiana in 15% and B. garinii in 15% of the positive ticks (3% could not be determined). In one tick, a double infection with B. valaisiana type II and B. garinii OspA type V was detected. Female adult ticks had the highest infection rate (11.7%), followed by nymphs (4.5%) and larvae (3.4%). The overall prevalence increased from spring (4.0%) to autumn (10.0%). Nevertheless, the risk of infection was maximal in summer, because of a much higher infestation and consequently a higher absolute number of infected ticks. The predominance of B. afzelii probably results from its resistance against human serum. The unexpectedly low total prevalence is possibly caused by immune defence mechanisms (e.g. complement) effective against less resistant B. burgdorferi s.l. strains in the tick.

A potential role of seabirds in spreading Lyme disease (LB) spirochetes over
large spatial scales was suggested more than 10 years ago when Borrelia garinii
was observed in marine birds of both hemispheres. Since then, there have been
few studies examining the diversity of Borrelia spp. circulating in seabirds, or
the potential interaction between terrestrial and marine disease cycles. To
explore these aspects, we tested 402 Ixodes uriae ticks collected from five
colonial seabird species by amplification of the flaB gene. Both the average
prevalence (26.0%+/-3.9) and diversity of LB spirochetes was high. Phylogenetic
analyses grouped marine isolates in two main clades: one associated with B.
garinii and another with B. lusitaniae, a genospecies typically associated with
lizards. One sequence also clustered most closely with B. burgdorferi sensu
stricto. Prevalence in ticks varied both among seabird species within colonies
and among colonies. However, there was no clear association between different
Borrelia isolates and a given seabird host species.

**Our findings indicate that LB spirochetes circulating in the marine system
are more diverse than previously described and support the hypothesis that
seabirds may be an important component in the global epidemiology and
evolution of Lyme disease.**

Future work should help determine the extent to which isolates are
shared between marine and terrestrial systems.

The aim of the study was to determine the infection level of adult forms and larvae of ticks and mosquitoes with Borrelia burgdorferi in the forested areas of Szczecin. A total of 1699 ticks Ixodes ricinus, including 1422 nymphs, 277 adult forms and 2862 mosquito females representing the genera Aedes (89.6%) and Culex (10.4%) were collected between the years 2004 and 2005. A further 3746 larvae and 1596 pupae of Culex pipiens pipiens were colleted from water bodies.

Borrelia burgdorferi s. l. was detected in the arthropods by the method of indirect immunofluorescence assay (IFA). A positive immunological reaction was detected in 16.6% of the adult forms and in 16.5% of the nymphs of Ixodes ricinus. Spirochetes were also detected in 1.7% of mosquito females, 3.2% of larvae and in 1.6% of pupae of Culex pipiens pipiens. The results of the present study confirm that contact with ticks constitutes the main risk of contracting Lyme disease, although mosquitoes play a role as vectors as well.

A total of 662 samples (winter period: 469; summer period: 193 specimens) of female mosquitoes of the genus Culex, Aedes and Anopheles were collected during the period March 2000-April 2001 from the locality of Vysoke Myto (Eastern Bohemia, Czech Republic). They were examined by dark field microscopy for the presence of spirochetes. The motile spirochetes were observed in 4.2 % of all species of investigated mosquitoes. One spirochetal strain out of the 8 isolation attempts (BRZ14) was obtained (cultivation rate was 12.5 %) and the spirochetal strain was then successfully cultivated and identified using PCR for the presence of Borrelia burgdorferi s.l., and subsequently with the RFLP as genomospecies Borrelia afzelii. This strain was derived from overwintering Culex (Culex) pipiens biotype molestus female mosquitoe. This is apparently one of the sporadic cases of the occurrence of pathogenic borreliae in haematophagous arthropods, other than Ixodes ricinus complex ticks.

The aim of the study was to determine the level of infection in mosquitoes with spirochetes Borrelia burgdorferi sensu lato in the woody areas of Szczecin. The mosquitoes were collected from May to September 2003. The spirochetes, Borrelia burgdorferi s. l., present in mosquitoes were detected in mosquitoes with indirect immunofluorescence assay (IFA) using rabbit anti-Borrelia burgdorferi antibodies and goat anti-rabbit IgG marked with fluorescein isocyanate (FITC). A total of 1557 females and 58 males were collected. They represented the genera Aedes (63%) and Culex (37%). The infection level of the mosquitoes from the area studied amounted to 1.7%. The results of the present study confirm the potential of these arthropods to spread Lyme borreliosis.

Aim of this study was to evaluate cellular fatty acid analysis for characterization of spirochetes. Strains were isolated from arthropods collected in South Moravia, Czech Republic. Fatty acid methyl esters (FAME) profile was determined for five Borrelia burgdorferi sensu lato (s.l.) strains isolated from Ixodes ricinus ticks, one "Spironema culicis" strain recovered from mosquito Culex pipiens and seven spirochetal strains (not identified yet) isolated from mosquitoes and blackflies. Analysis was performed using a gas chromatography column in conjunction with Microbial Identification System Sherlock (MIDI Inc., Newark, DE, USA). Results obtained on the basis of cluster analysis of FAME profiles showed, that the B. burgdorferi sensu lato isolates could be well separated from other spirochetal isolates. We recommended method used in this study as a useful tool for preliminary identification of spirochetes isolated from ticks and dipterans.

After first finding Borrelia in the midgut of imago mosquitoes, we concentrated on the presence of Borrelia in mosquito development stages--larvae of the third or fourth instar. In the summer season in the years of 2000-2001 a total of 439 Culex (Culex) pipiens pipiens larvae were collected from a barrel of rainwater in the Obrany holiday area of Brno city (East Moravia, Czech Republic). The larvae midgut was observed under dark-field microscopy. Ten DFM positive samples (2.28%) were further analysed using the single-tube nested PCR method for the presence of flagellum DNA sequence specific for Borrelia burgdorferi sensu lato, of which 5 were positive. Borrelian positivity of Culex (C.) pipiens pipiens larvae was 1.14%. One spirochete isolated strain in BSK-H medium was obtained. PCR detection for borrelian DNA of the isolated strain was negative. From these results we can conclude that a low percentage Borreliae can be also found in mosquito larvae and are likely to survive into imago stage.

Colonial seabirds often breed in large aggregations. These individuals can be exposed to parasitism by the tick Ixodes uriae, but little is known about the circulation of pathogens carried by this ectoparasite, including Lyme disease Borrelia. Here we investigated the prevalence of antibodies (Ab) against Borrelia burgdorferi sensu lato in seabird species sampled at eight locations across the North Atlantic. Using enzyme-linked immunosorbent assay tests, we found that the prevalence of anti-Borrelia Ab in adult seabirds was 39.6% on average (over 444 individuals), but that it varied among colonies and species. Common guillemots showed higher seroprevalence (77.1%+/-5.9) than black-legged kittiwakes (18.6%+/-6.7) and Atlantic puffins (22.6%+/-6.3). Immunoblot-banding patterns of positive individuals, reflecting the variability of Borrelia antigens against which Ab were produced, also differed among locations and species, and did not tightly match the prevalence of Borrelia phylogroups previously identified in ticks collected from the same host individuals. These results represent the first report of the widespread prevalence of Ab against Borrelia within an assemblage of seabird species and demonstrate that Borrelia is an integrated aspect in the interaction between seabirds and ticks. More detailed studies on the dynamics of Borrelia within and among seabird species at different spatial scales will now be required to better understand the implications of this interaction for seabird ecology and the epidemiology of Lyme disease.

Borrelia spirochetes in bird-feeding ticks were studied in the Czech Republic.

During the post-breeding period (July - September 2005), 1,080 passerine birds were infested by 2,240 Ixodes ricinus subadult ticks. Borrelia garinii was detected in 22.2% ticks, B. valaisiana in 12.8%, B. afzelii in 1.6%, and B. burgdorferi s.s. in 0.3%. Analyzing infections in the context of blood meal volume and stage of ticks, we concluded that European blackbirds Turdus merula, song thrushes T. philomelos and great tits Parus major are capable of transmitting B. garinii; that juvenile blackbirds and song thrushes are prominent reservoirs for B. garinii spirochetes; that other passerine birds investigated play minor roles in transmitting B. garinii; and that the presence B. afzelii in ticks results from infection in former stage. While B. garinii transmission is thus associated with few passerine bird species, these have potential for distributing millions of Lyme disease spirochetes between synanthropic areas.

The effect of introduced large herbivores on the abundance of Ixodes ricinus and their Borrelia infections was studied in a natural woodland in the Netherlands. Oak and pine plots either ungrazed or grazed by cattle were selected. Ticks were collected weekly by blanket dragging. Borrelia infections were determined by PCR and RFLP. Rodent densities were estimated using mark-release recapture methods.

On occasion, the cattle were inspected for tick infestations. Meteorological data were recorded in each habitat. Significantly more ticks were collected in the ungrazed woodland than in the grazed woodland. Ungrazed oak habitat had higher tick densities than pine habitat, while in the grazed habitats tick densities were similar. Borrelia infections ranged from zero in larvae, 26% in nymphs to 33% in adult ticks and consisted of B. afzelii, B. burgdorferi s.s., B. garinii or B. valaisiana. Co-infections were found in five ticks. There was no effect of cattle on Borrelia infections in the ticks. In the ungrazed area Borrelia infections in nymphs were significantly higher in oak habitat than in pine habitat. More mice were captured in the ungrazed area and these had a significantly higher tick burden than mice from the grazed area. Tick burden on cattle was low. The results suggest that grazing has a negative effect on small rodents as well as on ticks, but not on Borrelia infections. Implications of these results for management of woodland reserves and risk of Lyme disease are discussed.

Department of Medical Clinic, Dermatology Division, School of Medical Sciences, State University of Campinas, Campinas, Sao Paulo, Brazil.
renatafmagalhaes@uol.com.br

Bartonella henselae is the agent of cat scratch disease and bacillary angiomatosis. Blood donors can be asymptomatic carriers of B. henselae and the risk for transmission by transfusion should be considered. The objective of this study was to demonstrate that B. henselae remains viable in red blood cell (RBC) units at the end of the storage period. Two RBC units were split into two portions. One portion was inoculated with B. henselae and the other was used as a control. All units were stored at 4 degrees C for 35 days. Aliquots were collected on a weekly basis for culture in a dish with chocolate agar, ideal for the cultivation of this agent. Samples were collected on days 1 and 35 and taken for culture in Bact/Alert R blood culture bottles. Aliquots taken simultaneously were fixed in Karnovsky's medium for subsequent electron microscopy evaluation.

The viability of B. henselae was demonstrated after a storage period of RBC units. These data reinforce the possibility of infection by transfusion of blood units collected from asymptomatic blood donors.

Paris battles invasion of Siberian chipmunk
Siberian chipmunks have invaded forests around Paris in their tens of thousands, many carrying the potentially lethal Lyme disease.

By Henry Samuel in Paris
Last Updated: 3:52PM BST 20 Oct 2008

Experts are advising people to steer clear of the rodents. Pest specialists are calling for a ban on the sale of Tamias Sibiricus, the Siberian cipmunk known in France as the Korean squirrel. The tiny - but bold - furry creature has five stripes running down its back and its population has exploded in woods all around the French capital - including Versailles.

Experts are advising people strolling in the city's parks and surrounding woodland to steer clear of the rodents, as many of them carry ticks which are infected with Lyme disease.

The chipmunk was first imported from east Asia in the 1970s but a few specimens clearly escaped or were released into the wild, and have successfully adapted to their new home, breeding like wildfire.

"It's impossible to know how many there are overall, but we estimate that at one single site in the southern suburbs - the forest of Sénart - there are several thousand of the chipmunks," said Julie Marmet, a researcher at the Natural History Museum in the French capital.

The inquisitive chipmunks spend most of their time foraging on the ground but like squirrels they flee to trees if threatened. The chipmunks have been placed on the list of the European Union's 100 most invasive species, but experts say it is too early to tell whether they will be as damaging to indigenous species as the grey squirrel in Britain.

The red squirrel population is still relatively intact in France, which is free from grey squirrels.

Scientists want sales of the prolific chipmunk banned as they fear it poses a health hazard to humans. Studies on the colony in the Sénart woods show that up to a third of the chipmunks carry the Borrelia virus which causes Lyme disease - a dangerous nervous condition transmitted by ticks that can be fatal if not treated early.

"The point is that the chipmunks are much less shy than other rodents, so they are more likely to come into contact with humans. We think it is a basic precaution that the animals should be banned from sale in pet shops," said Jean-Louis Chapuis, France's leading expert on the non-indigenous rodents.

Human granulocytic ehrlichiosis was first described in the United States, in the northern Midwest, in 1994.1 Human granulocytic ehrlichiosis is caused by an organism, still referred to as the agent of human granulocytic ehrlichiosis, that is similar to two animal pathogens, Ehrlichia phagocytophila and E. equi.2,3,4 Transmission of human granulocytic ehrlichiosis occurs through the bites of ixodes ticks, which are the arthropod vectors for Borrelia burgdorferi and Babesia microti.5,6 Human granulocytic ehrlichiosis is an acute, febrile, nonspecific illness that may be severe enough to cause hospitalization and even death, particularly in the elderly.1,7,8 We describe a case of human granulocytic ehrlichiosis that developed in a pregnant woman near term and was transmitted perinatally to her infant.Case Reports

Mother

A 35-year-old woman, 39 weeks pregnant, was admitted to the hospital on October 4, 1997, at the onset of uterine contractions. She had malaise and had been feverish earlier that day. She had had an episode of Lyme disease with erythema migrans and antibodies to B. burgdorferi 10 years earlier and had a history of urinary tract infections. She lived in a tick-infested area of Connecticut. She recalled finding ticks crawling on her one week before admission, but none had embedded themselves in her skin. On admission her temperature was 38.1°C. Laboratory studies were limited to a complete blood count, which revealed a white-cell count of 6300 per cubic millimeter (normal range, 4600 to 11,200), with 5 percent lymphocytes, 75 percent neutrophils, and 17 percent band forms; a hematocrit of 35 percent (normal range, 36.4 to 45.8 ); and a platelet count of 168,000 per cubic millimeter (normal range, 160,000 to 410,000). Urinalysis revealed no white cells. On the following day, the patient had a normal vaginal delivery without complications. Fetal-scalp monitoring was not used. Therapy with clindamycin and gentamicin was begun because of persistent fever. The physical examination was normal, and there were no rashes. A chest roentgenogram, sinus radiographs, and pelvic magnetic resonance images were interpreted as normal. Two sets of routine blood cultures obtained that day were negative. A cervical culture was positive for group B streptococcus.

On the day after delivery, the woman's temperature rose to 40.6°C and she reported chills, malaise, fever, and myalgias. The results of a physical examination again were unremarkable. On that day, her white-cell count was 5600 per cubic millimeter, her hematocrit was 32 percent, and her platelet count was 130,000 per cubic millimeter. Over the next four days, she remained febrile without an identified source of infection. However, she noted some gradual improvement in her systemic symptoms. On October 9, she had a white-cell count of 5000 per cubic millimeter with 20 percent lymphocytes and a platelet count of 98,000 per cubic millimeter. On October 10, her alkaline phosphatase level was 186 U per liter (normal range, 30 to 115), her aspartate aminotransferase level was 74 U per liter (normal range, 7 to 40), her alanine aminotransferase level was 68 U per liter (normal range, 7 to 40), and her lactate dehydrogenase level was 420 U per liter (normal range, 100 to 225). On October 9, a whole-blood sample obtained on October 7 and treated with acid?citrate?dextrose as an anticoagulant demonstrated DNA from the agent of human granulocytic ehrlichiosis on polymerase-chain-reaction (PCR) assay, and a buffy-coat preparation from October 9 revealed morulae in granulocytes. The following day, clindamycin and gentamicin therapy was discontinued, and doxycycline treatment (100 mg orally twice daily) was started. Within 24 hours the patient became afebrile. She was treated for five days and remained well eight months later. The agent of human granulocytic ehrlichiosis was identified in a culture three days after inoculation with a sample taken on October 10, before the beginning of doxycycline therapy.

Sequential serologic indirect-immunofluorescence assays for the agent of human granulocytic ehrlichiosis demonstrated an increase in the antibody titer from 1:80 on October 10 to 1:2560 or more on October 17. An enzyme-linked immunosorbent assay (ELISA) did not detect IgG or IgM antibodies to B. burgdorferi in serum obtained on October 10. However, an ELISA of serum obtained on October 17 was positive for both IgG and IgM antibodies, and Western blotting was positive for IgM, with bands of 93, 66, 41, 39, 35, 29, and 24 kd (that at 24 kd indicates the presence of outer surface protein C [OspC]). B. burgdorferi was not detected by PCR in whole blood obtained on October 10.

Infant

On October 5, a 3000-g girl was born to the patient. The Apgar scores were 8 and 9 at one and five minutes, respectively. Breast-feeding was stopped after 24 hours because of the mother's illness and was not resumed because of the mother's use of doxycycline. The expressed breast milk was discarded. At six days of life, the infant was discharged with her mother. The next day the mother noted that the baby felt warm but fed well. On October 13, the child's ninth day of life, she had a temperature of 39.4°C and was referred for admission.

The physical examination on admission was normal. Treatment with ampicillin and gentamicin was begun after a workup for sepsis. A complete blood count revealed a white-cell count of 5200 per cubic millimeter, with 37 percent neutrophils, 12 percent band forms, and 39 percent lymphocytes, 10 percent monocytes, and 1 percent metamyelocytes; a hematocrit of 37.2 percent; and a platelet count of 92,000 per cubic millimeter. Liver enzyme levels, urine, and cerebrospinal fluid were normal. The cerebrospinal fluid was not examined for morulae. Because human granulocytic ehrlichiosis had been diagnosed in the mother, a buffy-coat smear of the infant's blood was examined, revealing morulae in 23 percent of granulocytes (Figure 1). A retrospective PCR analysis of the infant's blood spot obtained at birth did not detect DNA of the agent of human granulocytic ehrlichiosis.

View larger version (140K):[in this window][in a new window] Figure 1. Photomicrograph of a Buffy-Coat Smear Stained with Wright's Stain, Showing the Infant's Granulocytes Infected with the Agent of Human Granulocytic Ehrlichiosis (x1000).Arrows indicate morulae.

After the benefits and risks had been considered, intravenous doxycycline treatment (5 mg per kilogram of body weight per day, divided into two doses) was begun. Within 24 hours, the baby's body temperature returned to normal and her condition was clinically improved. The platelet count and neutrophil count reached nadirs of 66,000 and 990 per cubic millimeter, respectively. Two days after the initiation of treatment, the platelet count rose to 194,000 per cubic millimeter, and 7 percent atypical lymphocytes were found on a peripheral-blood smear. Blood obtained on the day of admission for cultivation of the agent of human granulocytic ehrlichiosis was positive two days after inoculation. A PCR analysis performed on blood obtained at admission also revealed DNA of the agent of human granulocytic ehrlichiosis. Routine blood, urine, and cerebrospinal fluid cultures performed at admission were negative. The child was discharged in good health after five days of intravenous doxycycline. At that time, no morulae were visualized on buffy-coat smears.

An EDTA-treated sample of whole blood obtained from the infant on October 10, five days after birth and three days before her illness, was positive for the agent of human granulocytic ehrlichiosis according to PCR, but negative for morulae. A specimen from that date could not be cultured for the agent of human granulocytic ehrlichiosis because of bacterial contamination. Serologic tests revealed no antibodies to B. burgdorferi on October 23, 18 days after birth, but were positive, with a titer of 1:320, for antibodies to the agent of human granulocytic ehrlichiosis. PCR analysis was negative for B. burgdorferi. The PCR products of cultured agents of human granulocytic ehrlichiosis from the mother and child had identical restriction-fragment?length polymorphisms.

Methods

Evaluation for Infection with the Agent of Human Granulocytic Ehrlichiosis

Buffy-coat smears were stained with Wright's stain, and 1000 granulocytes were examined at a magnification of 500 and 1000 for intragranulocytic morulae. PCR testing was performed on EDTA-treated whole blood to detect the agent of human granulocytic ehrlichiosis by the nested procedure of Sumner et al.,3 with the use of primers HS1/HS6 and HS43/HS45. Serologic analysis for antibodies to the agent of human granulocytic ehrlichiosis was performed with an indirect-immunofluorescence assay that used homologous and heterologous Westchester County isolates of the agent of human granulocytic ehrlichiosis cultured in HL-60 cells.8 HL-60 cell cultures were performed by adapting the techniques described by Goodman et al.9 and were evaluated by Wright's staining for the presence of morulae.8,9 The presence of the agent of human granulocytic ehrlichiosis in cultures was confirmed by PCR analysis and an indirect-immunofluorescence assay that used another patient's high-titer antiserum. Restriction-fragment?length polymorphism analysis of the agent of human granulocytic ehrlichiosis was carried out on a 332-bp fragment of the 16S?23S ribosomal DNA intergenic spacer. This region was amplified by species-specific PCR, the resulting product was digested with either HphI or DdeI, and the digests were analyzed by electrophoresis on 2.5 percent agarose gels.

Evaluation for Infection with B. burgdorferi

An ELISA (Wampole Laboratories, Cranbury, N.J.) was used to test for IgG and IgM antibodies to B. burgdorferi. Individual immunoblot assays for IgG and IgM antibodies to B. burgdorferi (MarDx Diagnostics, Carlsbad, Calif.) were performed and interpreted according to published criteria.10 PCR analysis to detect B. burgdorferi DNA in whole blood treated with an anticoagulant was performed as previously described.11

Discussion

Infection by the agent of human granulocytic ehrlichiosis is an increasing public health concern in the United States and Europe. The agent of human granulocytic ehrlichiosis replicates within granulocytes circulating in peripheral blood, and the case presented here demonstrates that perinatal transmission of human granulocytic ehrlichiosis can occur. The case also sheds some light on the manifestations of clinical human granulocytic ehrlichiosis in the neonatal period.

The mother was apparently infected with the agent of human granulocytic ehrlichiosis toward the end of pregnancy and gave birth to a normal infant. Whether infection with the agent of human granulocytic ehrlichiosis earlier in pregnancy would have had more severe sequelae for mother or child is not known. In sheep and cows, E. phagocytophila causes stillbirth or abortion.12,13 Recent experiments have shown that E. phagocytophila can be transmitted across the placenta in cows.14 Another ehrlichial species, E. risticii, causes abortion and is transmitted transplacentally in horses.15

Clinical disease caused by the agent of human granulocytic ehrlichiosis has rarely been reported in children.7 The reason for this is a matter for speculation. Young sheep and dogs infected with E. phagocytophila and an E. equi?like organism, respectively, have less severe clinical illness than older animals.16,17 However, the infant we describe had a clinical presentation and laboratory abnormalities similar to those found among infected adults,7,8 and a very high percentage of this infant's granulocytes were infected with the agent of human granulocytic ehrlichiosis.

The route of infection of the infant could not be determined. The timing of the onset of illness is consistent with all three potential routes of infection (intrauterine, intrapartum, or through breast-feeding). Although it is tempting to speculate that the agent of human granulocytic ehrlichiosis was transmitted transplacentally, this could not be proved, because the umbilical-cord blood and the placenta had been discarded by the time the infant became ill. The sensitivity of PCR analysis of the dried blood from the neonatal blood-spot card is not known, so the negative results cannot be interpreted with confidence. The frozen expressed breast milk was also discarded before we could test it.

Although we suspect transplacental transmission as the route of infection of the infant, we cannot exclude the possibility that secretions containing blood from the birth canal were introduced into the baby through minor skin abrasions or during suctioning of the respiratory tract. E. phagocytophila has been found in leukocytes from the milk of cows infected with this organism.18 However, we believe that transmission of the agent of human granulocytic ehrlichiosis in breast milk is not a likely route of infection in this case because of the small amount of colostrum produced on the first (and only) day of breast-feeding.

The infant's rapid response to the short course of doxycycline is reassuring. However, even short courses of tetracyclines in pregnant women can lead to tooth discoloration in their children. In this case, given the illness of the child and the lack of clinical data on other antibiotics for the treatment of human granulocytic ehrlichiosis, the benefits of using doxycycline appeared to outweigh the risks. Trovafloxacin (a quinolone antibiotic) and rifampin have in vitro activity against the agent of human granulocytic ehrlichiosis19 but have not been tested in patients with ehrlichiosis. Long-term follow-up of the infant will be required to determine whether human granulocytic ehrlichiosis causes neurodevelopmental problems like those described in human monocytic ehrlichiosis infection of young children.20

The meaning of the mother's positive Western blot assay for IgM antibodies to B. burgdorferi is uncertain. She had no erythema migrans rash. Coinfection with the agent of human granulocytic ehrlichiosis and B. burgdorferi has been proved by culture of both organisms from samples taken simultaneously from a patient.21 However, antibodies alone cannot be used for the diagnosis of Lyme disease in patients acutely infected with the agent of human granulocytic ehrlichiosis, because the production of cross-reactive antibodies is likely.22[b] In this case, the mother's history of Lyme disease might have increased the likelihood that nonspecific B. burgdorferi?reactive antibodies would be produced in the setting of acute disease.

Transplacental transmission of B. burgdorferi, which may have devastating consequences in early pregnancy, has been well described.23 We argue that the agent of human granulocytic ehrlichiosis may also be transmitted by this route. This possibility raises the question of how to treat pregnant women who have had tick bites. In the general (nonpregnant) population, prophylactic antibiotics should not be prescribed routinely after tick bites.24 However, some authorities suggest prophylaxis for pregnant women with tick bites.25 More data are needed to determine the timing and choice of antibiotic for the treatment of pregnant women and newborn infants exposed to or infected by the agent of human granulocytic ehrlichiosis in areas where this disease is endemic.

Supported in part by grants from the Westchester County Department of Health (CMC-2502, to Dr. Horowitz, and HLT-27017, HLT-27018, and HLT-27019, to Dr. Aguero-Rosenfeld), the National Institute of Arthritis and Musculoskeletal and Skin Diseases (RO1-AR41511, to Dr. Schwartz), and the Centers for Disease Control and Prevention (U50/CCU213698-01-1, to Drs. Wong and Chu).

We are indebted to Fatemeh Kalantarpour, Mehdi Baluch, and Shoba Varde for their technical help in performing assays for the agent of human granulocytic ehrlichiosis and B. burgdorferi and to Kris Keenan for helping in the care of the child.

Source Information

From the Department of Medicine, Division of Infectious Diseases (H.W.H.), and Department of Pathology (M.A.-R.), Westchester County Medical Center and New York Medical College, and the Department of Biochemistry and Molecular Biology, New York Medical College (D.L., I.S.) Valhalla, N.Y.; the Department of Pediatrics, Section of Neonatology (E.K., E.K.J.), and the Department of Pathology (R.K.), Danbury Hospital, Danbury, Conn.; the Department of Medicine, United Hospital, Port Chester, N.Y. (S.H.); and the Wadsworth Center, New York State Department of Health, Albany (S.J.W., F.C.).

http://www.vetscite.org/publish/items/005155/index.html 21 April 2009 Pets may become latest victims of climate change Pets are normally sheltered from the harsh realities of wild living. But across Europe, increasing temperatures will expose pets to new infectious diseases spread by ticks, fleas and mosquitoes, according to new research. Tick populations already appear to be increasing with the change in seasons. As winters become milder, ticks are becoming active all year round. The European dog tick is transmitting a malaria-like disease, canine babesiosis, into countries where it was once rare including Belgium, Germany, Poland and the Netherlands.Meanwhile, Ixodes ticks are living at greater densities across Europe, increasing their risk of passing tick-borne encephalitis to horses and dogs. Cat flea typhus, still a rare disease, may also become more common in both cats and dogs, according to Frederic Beugnet of Merial Animal Health in Lyon, France.In a separate paper, Claudio Genchi of the University of Milan, Italy, has found that dogs in central Europe will increasingly become vulnerable to the roundworm dirofilaria, spread by mosquitoes, as summer temperatures climb high enough for the parasite to incubate in its fly host.Susan Shaw and colleagues at the University of Bristol, UK, have also found a significant reservoir of canine leishmaniosis in dogs living in the southern UK. If climate change allows sandflies to spread into the country, there is a real danger the disease could spread, they warn.Source: Veterinary Parasitology New ScientistApril 21, 2009

We report a case of Lyme disease with clinical features resembling those described from brown recluse spider bites. The most striking manifestation was a necrotic skin wound. Brown recluse spider bites may be overdiagnosed in some geographic regions. Tick bite and infection with Borrelia burgdorferi should be considered in the differential diagnosis of necrotic arachnidism in regions endemic for Lyme disease.

"Tick bites, clinical symptoms of Lyme borreliosis, and Borrelia antibody responses in Finnish army recruits training in an endemic region during summer.

Tick bites, associated clinical symptoms, and antibodies against Borrelia burgdorferi were investigated in 77 Finnish army recruits training during summer in an endemic region and 50 control recruits serving outside the habitat of ticks. During a follow-up of 6 months, 26.9% of the study recruits reported tick bites. None gave a history of erythema migrans. Five study recruits and none in the control group had a combination of two or three nonspecific symptoms compatible with Lyme borreliosis. Three of these five study recruits had been bitten by several ticks. Thirteen (16.9%) study recruits and two (4.0%) control recruits had positive antibody levels against B. burgdorferi in their first serum specimens. No significant change in immunoglobulin G antibody levels was seen between the first and second specimens. Immunoglobulin M antibody levels increased in the sera of 13 (11.9%) study recruits and 1 (3.0%) control recruit. We conclude that recruits training in tick habitats are at high risk of tick bites. All recruits starting their military training in endemic areas should be taught to recognize and remove ticks and advised to wear protective clothing.

Abstract
To investigate the involvement of lizard species in the natural cycle of Borrelia burgdorferi sensu lato (s.l.) in Hungary, a total of 186 reptiles belonging to three species-126 green lizards (Lacerta viridis), 40 Balkan wall lizards (Podarcis taurica), and 20 sand lizards (Lacerta agilis)-were captured in 2007 and 2008. All ticks removed from the lizards were Ixodes ricinus, either larvae (324/472; 68.6%) or nymphs (148/472; 31.4%). More than half (66/126; 52.4%) of L. viridis individuals were infested, and the prevalence of tick infestation on both the other two species was 35% each. All 472 I. ricinus ticks and tissue samples collected from 134 collar scales and 62 toe clips of lizards were further analyzed for the presence of B. burgdorferi s.l. with polymerase chain reaction. The amplification of B. burgdorferi s.l. DNA was successful in 8% (n = 92) of L. viridis, 9% (n = 32) of P. taurica, and 10% (n = 10) of L. agilis tissue samples. Restriction fragment length polymorphism genotyping identified the species Borrelia lusitaniae in all tested lizard samples. Prevalence of B. burgdorferi s.l. in ticks collected from L. viridis, P. taurica, and L. agilis was 8%, 2%, and 0%, respectively. Most of the infected ticks carried B. lusitaniae (74% of genotyped positives); however, Borrelia afzelii (5%) and B. burgdorferi sensu stricto (21%) were detected in ticks removed from green lizards and Balkan wall lizards, respectively. We conclude that lizards, particularly L. viridis, can be important hosts for I. ricinus larvae and nymphs; thus, they can be regarded as reservoirs of these important pathogen vectors. The role of green lizards has been confirmed, and the implication of Balkan wall lizards is suggested in the natural cycle of B. lusitaniae at our study site.

BACKGROUND: Babesia microti, the primary cause of human babesiosis in the UnitedStates, is an intraerythrocytic parasite endemic to the Northeast and upperMidwest. Published studies indicate that B. microti increasingly poses a bloodsafety risk. The American Red Cross Hemovigilance Program herein describes thedonor and recipient characteristics of suspected transfusion-transmitted B.microti cases reported between 2005 and 2007.

STUDY DESIGN AND METHODS:Suspected transfusion-transmitted Babesia infections were reported bytransfusion services or were discovered through recipient-tracing investigationsof prior donations from donors with a positive test for B. microti in aserologic study. Follow-up samples from involved donors were tested byBabesia-specific immunofluorescence assay, Western blot, and/or real-timepolymerase chain reaction analysis. RESULTS: Eighteen definite or probable B.microti infections, including five fatalities, were identified in transfusionrecipients, 16 from hospital-reported cases and two through serologic lookbackstudies. Thirteen recipients were 61 to 84 years old and two were 2 years old oryounger. Two recipients had sickle cell disease and four were known to beasplenic, including one with sickle cell disease. Seventeen antibody-positivedonors were implicated; 11 (65%) were residents in Babesia-endemic areas, whilefour (24%) nonresident donors had a history of travel to endemic areas.

CONCLUSIONS: Transfusion-transmitted B. microti can be a significant cause oftransfusion-related morbidity and mortality, especially in infant, elderly, andasplenic blood recipients. These data demonstrate the need for interventions, inboth endemic and nonendemic areas of the United States, to reduce patient risk. http://eutils.ncbi.nlm.nih.gov/entrez/e ... rlinksPMID: 19624607 [PubMed - as supplied by publisher]

In this study, larvae (1,179 ex.) of mosquito genera Culex were examined for the
presence of spirochaetes by Dark Field Microscopy (DFM) at the locality of
Blansko (Czech Republic) in of 2004-2008.
DFM spirochaete positive samples (25.4%) were investigated by nested PCR; only 4 samples were positive for the presence of Borrelia burgdorferi sensu lato, which is 0.3 % of the total examined samples.

*We can conclude that only a low percentage of pathogenic borreliae are presented in mosquito larvae*, **while the spirochaete of undefined genera infect larvae in high amounts**.

"A bacterial infection typically spread by fleas, lice and biting flies could be more prevalent than many think, and may have been transmitted from a mother to her children at birth, scientists from N.C. State University say.

Dr. Edward Breitschwerdt, an infectious disease veterinarian and one of the world's leading researchers of bacteria called Bartonella, has for the first time documented evidence that the pathogen may have been passed between family members.

Although more studies are needed to back up his findings, Breitschwerdt and colleagues describe the case of a mother and father who began battling chronic aches, fatigues and other symptoms soon after they were married. When their twins were born in 1998, the daughter died after nine days from a heart defect, and the son developed chronic health problems.

Using tissue from the daughter's autopsy and blood from the surviving family members, Breitschwerdt's team discovered that the entire family was infected with the same species of Bartonella bacteria, despite having no shared exposures to flea or lice infestations. Bartonella is known to causes such illnesses as trench fever and cat scratch disease, and it is increasingly suspected of triggering a variety of aches and inflammations that doctors have been unable to diagnose.

"I think we have stumbled across something that is of monumental medical importance," said Breitschwerdt, whose findings were published recently in the Journal of Clinical Microbiology.

Proving the mother-child transmission could be difficult, however. Little funding is available for such research because the bacteria are still not considered a major source of human disease.

Dr. Michael Kosoy, who heads the Bartonella laboratory for the Centers for Disease Control and Prevention in Fort Collins, Colo., said scientists are only beginning to build evidence that Bartonella infections may be more common than previously thought.

"Bartonella are circulated around the world in many animals, but there are different Bartonella species, and the question is how can they be transmitted to humans?" Kosoy said, noting that most known cases have been transmitted from biting insects. He said the NCSU findings about the potential family transmission are compelling but inconclusive.

Dozens of strains

At least 26 strains of Bartonella have been named worldwide, and the list is growing. The most notorious Bartonella infection is cat scratch disease, a fever illness passed to humans from flea-infected cats. Fleas are the primary hosts, and they spread the bacteria in their feces.

Other Bartonella strains spread more serious diseases. Kosoy is studying how often heart inflammation is caused by a Bartonella that thrives among rat fleas in Thailand. He has already established that about 25 percent of unexplained fever illnesses among a group of patients there was caused by Bartonella .

"This is not limited to cat scratch," Kosoy said. "That's just the tip of the iceberg."

Breitschwerdt said he thinks the bacteria may be the hidden cause behind a host of chronic symptoms - muscle aches, neurological problems, fatigue, arthritis - that defy diagnosis.

About two years ago, Breitschwerdt began testing blood samples from a doctor in Maryland, who was curious whether Bartonella infections might be causing problems for some of his patients.

"There are lab tests showing inflammation," but no discernible cause, said Dr. Robert Mozayeni, a Yale-educated rheumatologist who practices in Rockville, Md.

Mozayeni contacted Breitschwerdt and his NCSU colleague, Ricardo Maggi, who together developed a more sensitive test for Bartonella. Routine blood tests fail to detect Bartonella because they search for antibodies that the body is slow to produce.

Instead, Breitschwerdt and Maggi figured out how to cultivate the bacteria in the laboratory from blood samples of infected people. They founded a company called Galaxy Diagnostics to handle the laboratory volume.

"I suspect this is going to be one of the causes of rheumatoid arthritis and a few other things, but it's too speculative right now to say," Mozayeni said.

Human testing

More studies are needed, and Mozayeni has joined Breitschwerdt and Maggi in the diagnostic company to oversee human testing.

"Certainly, the prevalence of Bartonella infection in people with chronic illness is higher than I would have ever guessed, but we still don't know what that means," Breitschwerdt said.

Among the biggest unknowns is how to treat people who have been infected. The effectiveness of antibiotics depends on which strain of Bartonella is at work, and with so many strains, treatments can be hit or miss.

Breitschwerdt said the family in his most recent study declined to comment about their experience. He said they were having difficulty finding a doctor.

"It is very difficult to find a physician who wants to see someone with a chronic illness that is poorly defined," he said, adding that many such patients often think they have Lyme disease, a tick-borne bacterial infection with similar symptoms - and stigma. "With an unexplained illness, it becomes problematic."

The importance of established and emerging tick-borne pathogens in Central and
Northern Europe is steadily increasing. In 2007 we collected Ixodes ricinus
ticks feeding on birds (N=211) and rodents (N=273), but also host-seeking stages
(N=196) in a habitat in Central Germany. In order to find out more about their
natural transmission cycles, the ticks were tested for the presence of Lyme
disease borreliae, Anaplasma phagocytophilum, spotted fever group (SFG)
rickettsiae, Francisella tularensis, and babesiae. Altogether, 20.1% of 680
examinded ticks carried at least one pathogen. Bird-feeding ticks were more
frequently infected with Borrelia spp. (15.2%) and A. phagocytophilum (3.2%)
than rodent-feeding (2.6%; 1.1%) or questing ticks (5.1%; 0%). Babesia spp.
showed higher prevalences in ticks parasitizing on birds (13.2%) and
host-seeking ticks (10.7%) whereas ticks from small mammals were less frequently
infected (6.6%). SFG rickettsiae and F. tularensis were also found in ticks
collected off birds (2.1%; 1.2%), rodents (1.8%; 1.5%) and vegetation (4.1%;
1.6%). Various combinations of co-infections occurred in 10.9% of all positive
ticks, indicating interaction of transmission cycles.
Our results suggest that birds are not only important reservoirs for several pathogens but also act as vehicles for infected ticks and might therefore play a key role in the dispersal of tick-borne diseases.

Med Vet Entomol. 2010 Sep 26; [Epub ahead of print]
Established and emerging pathogens in Ixodes ricinus ticks collected from birds
on a conservation island in the Baltic Sea.

Franke J, Meier F, Moldenhauer A, Straube E, Dorn W, Hildebrandt A.

Department of Food and Environmental Hygiene, Institute of Nutrition, Friedrich
Schiller University of Jena, Jena, Germany Medical University Laboratories,
Institute of Medical Microbiology, Friedrich Schiller University of Jena, Jena,
Germany.

Tick-borne pathogens such as Lyme borreliosis spirochaetes, Anaplasma
phagocytophilum, Rickettsia spp. and Babesia spp. cause a great variety of
diseases in animals and humans. Although their importance with respect to
emerging human diseases is increasing, many issues about their ecology are still
unclear. In spring 2007, 191 Ixodes ricinus (Acari: Ixodidae) ticks were
collected from 99 birds of 11 species on a bird conservation island in the
Baltic Sea in order to test them for Borrelia spp., A. phagocytophilum,
Rickettsia spp. and Babesia spp. infections. Identification of the pathogens was
performed by polymerase chain reaction (PCR), restriction fragment length
polymorphism and sequence analysis. The majority of birds with ticks testing
positive were European robins and thrushes. Borrelia DNA was detected in 14.1%,
A. phagocytophilum in 2.6%, rickettsiae in 7.3% and Babesia spp. in 4.7% of the
ticks. Co-infections with different pathogens occurred in six ticks (3.1%). The
fact that 11 ticks (five larvae, six nymphs) were infected with Borrelia afzelii
suggests that birds may, contrary to current opinion, serve as reservoir hosts
for this species. Among rickettsial infections, we identified Rickettsia
monacensis and Rickettsia helvetica. As we detected five Rickettsia spp.
positive larvae and two birds carried more than one infected tick, transmission
of those pathogens from birds to ticks appears possible.
Further characterization of Babesia infections revealed Babesia divergens and Babesia microti. The occurrence of Babesia spp. in a total of five larvae suggests that birds may be able to infect ticks, at least with Ba. microti, a species considered not to be transmitted transovarially in ticks. (c) 2010 The Authors. Medical and Veterinary Entomology (c) 2010 The Royal Entomological Society.

Abstract Borrelia burgdorferi, the bacterium that causes Lyme disease, is
transmitted to a susceptible host by Ixodes spp. tick bites. However, there is
uncertainty whether B. burgdorferi are shed from ticks by the fecal route. In
this study, B. burgdorferi&#x2013;infected ticks were fed on mice while confined
to a certain area of the skin by a capsule. During and after feeding, tick feces
were collected and placed in Barbour-Stoenner-Kelley (BSK)-II media for
cultivation and in sterile water for polymerase chain reaction (PCR) analysis.
Although none of the tested samples were culture positive for B. burgdorferi,
all but one of the fecal DNA samples from infected ticks were PCR positive.
These results indicated that B. burgdorferi were shed from feeding ticks during
defecation and suggest that the spirochetes did not remain viable once exposed
to the outside environment. This finding has important ramifications for
investigators interpreting B. burgdorferi&#x2013;specific PCR results when
conducting tick transmission experiments.

I am always amazed when I hear this question. How can anyone doubt that if spirochetes are in body fluids, which they are, that Lyme would be transmitted like any other STD, syphilis for instance.

We know that conventional health care is trying to shut down the possibility of chronic Lyme, and deny any possibility of sexual transmission. There is a very clear effort to present the public with the notion that there isn?t anything to worry about Lyme disease; that it is very easy to diagnose and cure.

I had one Harvard teaching Infectious Disease Specialist tell me that he didn?t believe in chronic Lyme, and he was clearly upset by what he felt was a ?panic? caused by hypochondriacs that are always looking for a technical reason for why they don?t feel good. He said, ??one year it was Candida?everyone thought they were infected with Candida?another year it is another hysteria, well now it is Lyme.?

Main stream medicine also denies the possibility of babies becoming infected in the womb if the mother is infected. Yet Dr. Alan MacDonald, a specialist in neonatal pathology back in the early eighties, studied and collected an immense collection of clear evidence showing babies ? some stillborn and some who died soon after birth ? all from Lyme disease.

The world wasn?t ready for the truth when he presented his evidence in Vienna at the ?Second International Symposium on Lyme Disease and Related Disorders.? The process he used for proving the presence of spirochetes was innovative and difficult for the scientists to accept.

It would require them to cross over into the dark and terrifying realm of a runaway infection that these slides pointed to. And for the most part, twenty years later, the medical main stream is still looking the other way and ignoring the thousands upon thousands of suffering infected patients who are crying for help.

The day is sure to come when this tick-borne infection is understood better. But in the meantime, there are very few doctors willing to risk the persecution and financial risk of treating Lyme disease. Doctors are being sued not by their unhappy patients, but by other doctors who choose to stay blind to the emerging truth.

Entrenched and conservative medical practitioners say Lyme cannot be transmitted sexually, however, cutting edge scientists are proving the opposite.Alzheimer?s Autism, MS, Rheumatoid Arthritis, Heart disease, psychiatric conditions and many other horrible diseases are being shown to involve Lyme disease as the cause in increasing cases.

So when you look to basic biology and study how infections are passed; it takes more faith to believe Lyme is not transmitted sexually than that it is

How can it be proven? That remains the problem.

Jenna Smith is the author of the novel ?The Goddess of Sumer? and numerous articles on health and fitness. Jenna?s goal is to help people discover cutting edge strategies for healing disease and attaining optimum health. Discover your miracle body at: http://www.Miracle-Body.com and http://www.LymeDiseaseResource.com

We determined if deer mice (Peromyscus maniculatus) could be infected by Borrelia burgdorferi and develop sufficient spirochetemia to infect larval Ixodes dammini. Ten P. maniculatus were infected orally with 0.05 ml phosphate buffered saline containing approximately 400 B. burgdorferi. On days 21 or 28 after infection (AI) larval I. dammini were fed on the deer mice. Each of the P. maniculatus developed antibodies (up to 7 log2) to B. burgdorferi and B. burgdorferi was isolated from the blood of 1 deer mouse on day 51 AI. Nymphs resulting from these larvae were then allowed to feed on 10 uninfected P. maniculatus. All 10 of these tick-infected P. maniculatus developed antibodies (up to 7 log2) to B. burgdorferi, and B. burgdorferi was isolated from the blood of 1 of the 10 P. maniculatus 15 days after tick feeding and from the pooled organs of another of the tick-infected P. maniculatus.

Six of the orally infected P. maniculatus developed clinical signs including ruffled hair coat, inappetence, reluctance to move, and lameness in the rear legs. All P. maniculatus tissues were grossly and histologically normal on necropsy.

These findings show that P. maniculatus are susceptible to oral infection and develop sufficient spirochetemias to infect I. dammini larvae.

Numerous reports exist of the transmission of zoonoses to humans during and after solid-organ and hematopoietic stem cell transplantation. Donor-derived infections of numerous etiologies, including West Nile virus infection, Chagas disease, toxoplasmosis, rabies, lymphocytic choriomeningitis virus infection, and infection due to Brucella species have been reported. Most zoonoses occur as a primary infection after transplantation, and immunocompromised patients are more likely to experience significant morbidity and mortality from these infections. Risks of zoonotic infection in the posttransplantation period could be reduced by patient education. Increased recognition of the risks of zoonoses, as well as the advent of molecular biolog-based testing, will potentially augment diagnostic aptitude. Documented zoonotic infection as it affects transplantation will be the primary focus of this review.

Zoonotic illnesses represent a significant risk to patients undergoing solid-organ transplantation (SOT) and hematopoietic stem cell transplantation (HSCT). Numerous reports exist of the transmission of zoonotic infection at the time of transplantation, either with the allograft or with blood products, as well as in the posttransplantation period, via the usual methods of transmission. The studies of zoonoses and transplantation-associated infectious diseases are evolving fields that are receiving increased recognition. Of the 1407 organisms that have been identified as human pathogens, 58% are zoonotic and are twice as likely as other pathogens to be in the ?emerging? category [1]. The population of immunocompromised hosts is also increasing; as the annual number of transplantations that are performed increases, transplant recipients are living longer [2], more-powerful immunosuppressive agents are being administered, and tools for the management of chronic graft-versus-host disease are improving.

Because numerous reviews of the effects of zoonoses on the general human population have been written, this review will primarily focus on documented zoonotic infection involving SOT and HSCT (excluding corneal and musculoskeletal grafting). Although much concern has been registered regarding the risk of zoonosis transmission with xenotransplantation (i.e., the transplantation of organs from animals to humans) [3], because this is not currently clinical practice, this will not be covered in this article. Cases described herein were found in reports in English-language journals via a search of the Medline database, using the search term ?transplant? along with the name of the genus or syndrome. The defining criteria for zoonoses that are covered herein, as previously defined [4], include pathogens that have a nonhuman vertebrate reservoir, entail transmission from animals to humans, and have a recognized infectious disease syndrome in susceptible humans. Infections that do not involve a nonhuman vertebrate intermediary, such as malaria and dengue fever, will not be included. Transmission may occur directly (via contact with infectious animals or their secretions), via a nonvertebrate vector, or indirectly (via food, water, or a shared environment) [5]. Significant zoonoses are covered in the text and in table 1, and rarer or les-commonly reported zoonoses are also included. Live viral vaccines are also discussed. In general, the incidence of zoonotic illness is not known to be higher in transplant recipients, although the related morbidity and mortality may be higher among this population.
Table 1
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Table 1

Zoonotic pathogens in transplant recipients.

Population shifts resulting from immigration and travel are occurring throughout the world. Approximately 10% of the population of the United States was born in a foreign country [126], and more Americans than ever before are traveling internationally. These are both factors that may augment the risk of donor-derived infections, particularly the more latent ones (i.e., infections due to Mycobacterium tuberculosis, Trypanosoma cruzi, parasites, and others). The rise in the number of SOTs performed in developing countries (some of which are performed in patients who return to industrialized countries after transplantation) may create another reservoir for unusual infections.

Immunocompromised patients may have atypical presentations of infectious diseases, and the diagnosis may be elusive, especially for some of the less common zoonoses. Many zoonotic illnesses involve specialized diagnostic tests. Transplant recipients may be slow to evolve a serologic response, which may delay or deter diagnosis, especially if their overall level of immunosuppression is high. The use of molecular biology-based tests, when available, may augment our diagnostic capacity in this population, similar to the contribution from nucleic acid amplification testing of the blood supply for West Nile virus (WNV) [127]. If donor-derived infection is a possibility, donor samples (e.g., serum, tissues, blood vessels, and autopsy specimens) should also be tested. Increasing recognition of the risks of zoonotic infection will potentially augment diagnostic aptitude.

Donor-derived infection. Zoonotic infection in the peritransplantation period can be transmitted via the organ or stem cell allograft, as well as through the transfusion of blood products. Both acute and latent infections (such as Chagas disease or toxoplasmosis) may be transmitted via an allograft. Risk factors for zoonotic illness may be overlooked during the standard screening process. Subclinical or atypical illness, such as was observed in the cases of lymphocytic choriomeningitis transmission [74] and rabies transmission [78, 79], may result in imperfect screening and subsequent transmission of infection. WNV infection [83], Chagas disease [124, 125], and toxoplasmosis [122, 123] have also been transmitted during SOT. WNV and Brucella infections have been transmitted in HSCT allografts. After transplantation, the clinical syndromes associated with these infections can be protean and may even be mistaken for transplant rejection (e.g., hepatitis after liver transplantation). When there is concern regarding donor-derived infection, both donor and recipient samples must be examined. Novel transmission should be promptly reported, because increased recognition may reduce subsequent infections.

Recipient-derived infection. Recipients in whom an infection is incubating at the time of transplantation and who subsequently experience profound immunosuppression in the peritransplantation period may develop severe infection. The majority of zoonoses are acquired after transplantation. Certain epidemiological risk factors increase the risk for acquisition of zoonoses, including occupational exposure (e.g., in veterinarians, pet store employees, farmers, slaughterhouse workers, landscapers, and forestry workers), pet ownership, hobbies (e.g., hunting), and travel. These exposures should be limited or possibly avoided, especially during the first 6 months after transplantation or other significant immunosuppression [128]. Perhaps more insidious are the risks that are more common and less obvious-that is, contaminated drinking water and food, walking in the woods or wading in the ocean, visiting a petting zoo, or exposure to any house pets.
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Pathogen-Specific Infection

The following sections include both donor- and recipient-derived infections, with a focus on the characteristics of specific pathogens, by category.
Bacterial Infection

Enteric infection. Bacterial enteric pathogens are common etiologies of foodborne and waterborne illnesses, and may represent contamination from farm animals or other vertebrate animals; they may also be directly transmitted from animals to humans. Transplant recipients have much higher rates of bacteremias due to Salmonellae species-ranging as high as 70%, compared with 3%?4% in normal hosts-and a higher risk of a metastasic focus of infection [129, 130]; this is true for other enteric pathogens, as well. Campylobacter species are another group of common zoonotic enteric pathogens in patients who undergo SOT or HSCT. As in other immunocompromised hosts, transplant recipients may have trouble completely clearing a Campylobacter jejuni infection [131]. Asymptomatic, prolonged Campylobacter species bacteremia in the peri-HSCT period has been documented [132]. Yersinia species can also cause invasive disease with bacteremia in SOT [62, 63]. Noncholera Vibrio species can cause fulminant illness in transplant recipients, with gastroenteritis, bacteremia, or skin and soft-tissue infections [57?60]; environmental flooding increases the risk of illness due to Vibrio species, as was observed after Hurricane Katrina [133]. Recent increases in drug-resistant bacteria, sometimes related to the use of antibiotics in animal feed (such as with multidrug-resistant Salmonella species), are especially concerning.

Pulmonary infection. Bordetella bronchiseptica, the etiologic agent of ?kennel cough? in dogs, has caused serious respiratory illness in patients who undergo pediatric lung transplantation [17], heart transplantation [18], and HSCT [19, 20]. Several of these case patients had pet dogs. The?kennel cough? live vaccine, which contains a mixture of parainfluenza virus and B. bronchiseptica, has the potential to cause human B. bronchiseptica infection [134]. Rhodococcus equi has been increasingly documented as a pulmonary pathogen in transplant recipients [135], as well as an agent of unusual infection, including cerebral infection in a heart transplant recipient [52], pericarditis in a kidney transplant recipient [136], and vertebral osteomyelitis in a liver transplant recipient [53]. Cases of tularemia (due to Francisella tularensis infection) after both HSCT and renal transplantation have been reported [33,34?35].

Bartonella henselae infection has been described after heart [9] and kidney transplantation [10,11,12?13], with variations in the manifestation of infection that include hemophagocytosis [137], closely associated acute allograft rejection [10], peliosis hepatis [138], peliosis hepatitis and hepatorenal syndrome [139], pulmonary nodules [140], and osteomyelitis [141]. Brucella species infection has been reported after kidney transplantation [21, 22] and as a donor-derived infection during HSCT [23], mostly in areas of endemicity. Live, attenuated Brucella animal vaccine has been linked to human disease and has the potential to cause disease in immunocompromised hosts [134]. Listeria monocytogenes infection has been well described in patients who undergo SOT and HSCT [37, 38, 41], including the rare clinical manifestations of tricuspid valve endocarditis with septic pulmonary emboli [39], epididymitis and orchitis [40], and skin infection with cerebritis and hemophagocytosis [42]. A small number of cases of human tuberculosis are due to Mycobacterium bovis infection, also known as zoonotic tuberculosis-an opportunistic infection in immunocompromised hosts [142]; M. bovis infection of the urinary tract has been documented after kidney transplantation [43].

Skin and soft-tissue infection. Numerous reports exist of Mycobacterium marinum infection in patients who undergo SOT, sometimes after a patient's exposure to fish [45,46,47?48]. Erysipelothrix species-related endocarditis that occurs after aquarium contact has been reported in a kidney transplant recipient [21]. Capnocytophaga species infections are usually caused by the organisms that are found in the oral flora of immunosuppressed hosts [143, 144] and not by the zoonotic species (i.e., Capnocytophaga canimorsus and Capnocytophaga cynodegmi, which have not been reported in transplant recipients); similarly, Pasturella species infections have not been reported in this population.
Fungal Infection

Cryptococcus species is the third most common cause of invasive fungal infection in organ transplant recipients after Candida species and Aspergillus species [64]. Birds and their droppings are the most commonly perceived risk. A 72-year-old kidney transplant recipient who owned a pet cockatoo developed cryptococcal meningitis that was believed to be acquired from the cockatoo, because isolates obtained from the patient and the bird had identical biochemical profiles, the same monoclonal antibody immunofluorescence patterns, and indistinguishable patterns on RFLP analysis and karyotyping [65]. Some authors suggest that immunocompromised hosts should not keep cockatoos, given their association with cryptococcosis [145].

Sporotrichosis due to Sporothrix schenckii infection can be connected to animal contact, especially contact with cats [146], and has caused severe, recurrent disease in a kidney transplant recipient [71]. Dermatophytes are common in both regular and exotic animals and can cause both superficial and invasive disease in humans [145, 147]. Trichophyton mentagrophytes, a zoonotic dermatophyte, was the most common superficial dermatophyte observed after kidney transplantation in 1 series [67]. The zoonotic dermatophyte Microsporum canis has caused invasive cutaneous infection after liver transplantation [69], relapsing tinea capitis after kidney transplantation [68], and dermatophytic granuloma with erythematous pustules and papules in a heart-lung transplant recipient [70].
Viral Infection

Viral zoonoses are numerous [4] and are common among the emerging zoonotic pathogens [148]; however, the vast majority have not been reported among transplant recipients. WNV infection is one of the more commonly reported viral zoonoses in transplant recipients. It may be donor derived, transfusion related, or normally acquired, and it carries a high morbidity and mortality. The risk of meningoencephalitis in a transplant recipient infected with WNV is estimated to be 40%-much higher than in normal hosts [84]. A recent case of WNV infection was confirmed in 3 of 4 recipients of organs transplanted from a single donor; 2 recipients subsequently experienced neuroinvasive disease, 1 recipient developed asymptomatic WNV infection, and a fourth recipient was apparently not infected [83]. Numerous additional reports exist of transmission following SOT and HSCT.

Rabies is rarely observed after SOT. In a recent case in Arkansas and Texas, 1 donor transmitted lethal rabies infection to 5 recipients [78], and in another case in Germany, 3 patients who underwent SOT developed neurological symptoms and died [79]. Live rabies vaccine for use in wildlife has caused human disease and presents a potential risk to transplant recipients who come in direct contact with it [134].

Yellow fever presents a risk to transplant recipients who are traveling to or whose donors are from areas of endemicity for the disease, although instances of infection in this manner have not been documented. Use of the live attenuated vaccine should be avoided in immunocompromised hosts [149]. Although a few immunosuppressed travelers have tolerated the vaccine (e.g., those in the early stages of HIV infection or who have a distant history of hematological malignancy [150,151?152]), complications, including death, have been reported [153].

A recent report documented the spread of lymphocytic choriomeningitis from 2 asymptomatic organ donors to 8 organ transplant recipients, 7 of whom died [74]. The severe acute respiratory syndrome (SARS) virus caused significant disease in patients who underwent HSCT and recipients of liver and kidney transplants [80,81?82]. Infection with parapoxvirus, the agent responsible for orf (ecthyma contagiosum) and milker's nodules, has been observed after HSCT (in these cases, the infection was transmitted by cows) [77] and kidney transplantation [75, 76].
Parasitic Infection

Numerous zoonotic parasites have been shown to cause disease in transplant recipients. Bloodborne and organborne infection may be transmitted at the time of transplantation; enteric pathogens are less likely to be transmitted during the peritransplantation period, although this could potentially occur with intestine and liver transplantation. Depending on the location and circumstances, toxoplasmosis, babesiosis, Chagas disease, and leishmaniasis are among the more common parasite-related infections observed in transplant recipients.

Toxoplasma gondii infection can be caused by primary infection transmitted by an allograft, as well as by reactivation disease. T. gondii allograft transmission is classically associated with heart transplantation, in which case can persist as a latent infection in the myocardium, although it has been transmitted through transplantation of other organs. Among patients who undergo HSCT, toxoplasmosis occurs in 0.3%?7.6% of cases, with higher rates in countries where toxoplasmosis is more prevalent and among patients with graft-versus-host disease [122]. The use of trimethoprim-sulfamethoxazole for post-SOT prophylaxis has decreased the risk of toxoplasmosis [154, 155]. In a recent review of 52 noncardiac SOT-related cases of toxoplasmosis, 86% of patients developed disease within 90 days of transplantation; of these patients, 42% had primary infection, 21% had reactivation or reinfection, and 37% had cases that could not be determined [123]. Classically, non-allograft-associated transmission was linked to the ingestion of either uncooked or undercooked meat containing viable tissue cysts or oocysts from the feces of infected cats; a report of an outbreak of toxoplasmosis associated with unfiltered municipal drinking water contaminated by felid waste [156] reiterates the importance of clean drinking water for transplant recipients.

Babesiosis has caused severe disease with hemophagocytosis and pancytopenia in asplenic renal transplant recipients [86, 87]. Babesia species have been transmitted through peritransplantation blood transfusions [85, 88]; in the United States, there has been a sharp increase in the number of transfusion-transmitted infections of Babesia species [157], suggesting a potential for increased infection in this generally heavily transfusion-dependent population.

Trypanosoma cruzi, the etiologic agent of Chagas disease, has been transmitted during SOT [124, 125], as well as during blood transfusions [158,159?160], and infection can also reactivate after transplantation [161]. Although most commonly associated with heart transplantation, other organs (including liver, kidney, and pancreas) may transmit T. cruzi as well [124, 125]. In a recent survey of 404 deceased organ donors in Southern California, where 25% of organ donors are of Hispanic ethnicity, 6 donors (1.5%) were found to be initially reactive by EIA, and 1 donor (0.25%) was found to have confirmatory T. cruzi antibodies, suggesting a beneficial role for the screening of transplant donors [162].

Leishmania species cause significant disease in immunocompromised hosts and could theoretically be transmitted via an allograft or a blood transfusion [163, 164]. Visceral disease (kala azar) is the most common manifestation after SOT, with 57 cases reported in the literature [105]. Patients who undergo HSCT appear to be rarely affected [116]. Cutaneous leishmaniasis has been reported in a handful of cases of SOT, some with concomitant visceral involvement [106,107,108,109?110]; mucosal disease has also been reported infrequently [111,112,113,114?115].

Enteric parasites are more likely to cause disease after transplantation. Microsporidia can cause infection in immunocompromised hosts; the most commonly reported is Enterocytozoon bieneusi infection following a transplantation [117]. Pulmonary infection has been described following allogenic HSCT [118, 119], and disseminated disease was documented postmortem in a kidney-pancreas transplant recipient [117]. Cryptosporidium parvum infection is especially common in patients who undergo SOT in the developing world [95]. Cryptosporidium species can also cause biliary disease and may play a role in some cases of otherwise unexplained cholangiopathies in liver [93] and kidney [96] transplant recipients. Disseminated Cryptosporidium species disease and related death have been described in liver [94] and stem cell transplant [97] recipients. Severe alveolar echinococcosis of the liver due to Echinococcus species has been successfully cured by liver transplantation [98,99,100?101], although there are risks of extrahepatic infection and potential echinococcal dissemination. Heart transplantation has been successfully performed in a patient who had hepatic echinococcosis [102].
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Recommendations for Pet Owners

Companion animals provide numerous benefits, along with some zoonotic risk. Discussions about pet ownership should optimally occur prior to transplantation. Pets may enhance health and well being, and many people would welcome advice and support to enable them to reconcile or manage pet ownership [165]. Guidance in pet choice can decrease zoonotic risk [128, 166]. In general, mature pets from reputable sources provide lower zoonotic risk. Fish are the pets least likely to be associated with illness (especially if aquarium cleaning by the transplant recipient is avoided). Animals to avoid as pets include reptiles (lizards, snakes, and turtles), baby chicks and ducklings, and exotic pets (chinchillas and monkeys); contact with stray and wild animals should also be avoided [128]. The individual risk of acquiring an infection from an animal is hard to calculate, and little work has been done in this field. In a survey of adult cats in Colorado, 13% were found to harbor zoonotic intestinal pathogens [167], and 41% of kittens in New York harbored a zoonotic agent [168].

Careful handwashing after any animal contact is imperative. Routine veterinarian care, with frequent stool examination for parasites, administration of routine vaccines, and evaluation when an animal is sick (especially with diarrhea), can reduce the risks of pet ownership to a transplant recipient. Immunocompromised hosts should avoid direct contact with any live viral vaccines that are administered to their pets and animals [134]. In addition, contact with animal excreta or saliva should be avoided. Good quality animal food should be given (not raw eggs or meat), and animals should not drink toilet bowl water. Humans should avoid flea and tick bites, as well as animal-related scratches and bites. Because small children are more likely to be bitten by pets and are less likely to practice good hand hygiene, pet ownership should potentially be deferred for very young transplant recipients. Pet therapy should potentially be avoided in hospitalized patients during the immediate posttransplantation period, when the patient is most immunosuppressed. The US Centers for Disease Control and Prevention's report on ?Pets and Organ Transplant Patients? [70] provides both general and animal-specific guidelines.
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Conclusions

Zoonotic infections are increasingly being recognized in transplant recipients, likely because of a greater number of transplantations, improved diagnostic testing in transplant recipients, and an augmented recognition of zoonoses.With increases in such factors as immigration, foreign travel, and exotic pet ownership, there may also be increased exposures to both donors and recipients. Risk of donor-derived infection may be reduced by improving the screening of donors, both through analysis of exposure history and through better molecular biology-based diagnostic testing. Given the current diversity and extent of animal contact, travel, occupational experience, and vector contact, a careful exposure history should be systematically ascertained in all transplant donors (when possible) and recipients. Education of transplant recipients before and after transplantation regarding zoonotic risks may further decrease zoonotic infection in this population.

DOI: 10.1111/j.1365-3016.1995.tb00148.x
Summary. This report describes a cohort study of over 5000 infants and their mothers who participated in a cord blood serosurvey designed to examine the relationship between maternal exposure to Lyme disease and adverse pregnancy outcome. Based on serology and reported clinical history, mothers of infants in an endemic hospital cohort are 5 to 20 times more likely to have been exposed to B. burgdorferi as compared with mothers of infants in a control hospital cohort. The incidence of total congenital malformations was not significantly different in the endemic cohort compared with the control cohort, but the rate of cardiac malformations was significantly higher in the endemic cohort [odds ratio (OR) 2.40; 95% confidence interval (CI) 1.25,4.59] and the frequencies of certain minor malformations (haemangiomas, Polydactyly, and hydrocele), were significantly increased in the control group. Demographic variations could only account for differences in the frequency of Polydactyly. Within the endemic cohort, there were no differences in the rate of major or minor malformations or mean birthweight by category of possible maternal exposure to Lyme disease or cord blood serology. The disparity between observations at the population and individual levels requires further investigation. The absence of association at the individual level in the endemic area could be because of the small number of women who were actually exposed either in terms of serology or clinical history. The reason for the findings at the population level is not known but could be because of artifact or population differences.
http://onlinelibrary.wiley.com/doi/10.1 ... x/abstract

".........Studies have demonstrated that deerflies, horseflies, and mosquitos can carry the B. burgdorferi spirochete......."

".....in some parts of the United States, 100% of the examined ticks have been infected.............

"...Ticks species that are themselves infected to a higher degree will increase the reservoir pool among the different animal species and increase the likelihood that human beings will become infected.(21,22,28-33) Studies also indicate that birds can serve as hosts........and thus act as reservoirs for B. burgdorferi. The bird reservoir allows for long distance dispersal of the spirochete within and between continents.........Studies have demonstrated that deerflies, horseflies, and mosquitos can carry the B. burgdorferi spirochete....The percentage of infected flies increases in areas where I. dammini is present. Therefore, these flies are probably serving as secondary vectors....."[I. dammini was name the given to the deer tick I. scapularis-- for a short time --someone thought it was a new species]

"An additonal concern for clinicians is the possibility that the ..tick may simulataneously expose an individual to two organisms that can present with central nervous system symptoms. Cases have been reported in Wisconsin and New York of people with direct infection as well as antibodies against both the Babesia microti organism and the B. burgdorferi spirochete." (104-106)-------------------------------------------------------------------------------------------------------------------------------------

Abstract:Lyme disease is reported from all over the United States. Transmitted by deer ticks, mosquitoes, and deer flies, it affects numerous organ systems. All age groups are vulnerable to this disease and must be educated about early signs and symptoms to speed diagnosis and appropriate treatment.

"...DiscussionThis is the first report of B. burgdorferi in horse flies, deer flies andmosquitoes. ....the number of infected deer flies and horse flies varied with the species and sampling areas....Also, like mosquitoes and other biting insects, the blood- feeding behavior of female tabanids differs within and between species, and infection may be correlated , in part, with the quantities of blood ingested....... ......Serological studies of mammals and identification of B. burgdorferi have established that this agent is widely distributed within given habitats in the United States and that closely related strains exist in Europe. The presence of this bacterium in tabanids and mosquitoes increases the risk of Lyme disease in tick infested areas. these and other blood sucking arthropods should recieve further consideration in ecological and epidemiological studies of this disease and of related disorders. ..."

Members of 18 species of ticks, mosquitoes, horse flies, and deer flues were collected in southeastern Connecticut and tested by indirect fluorescent-antibody staining methods for Borrelia burgdorferi, the etiologic agent of Lyme disease. An infection rate of 36.2% (116 tested), recorded for immature Ixodes dammini, exceeded positivity values for all other arthropod species. Prevalence of infection for hematophagous insects ranged from 2.9% of 105 Hybomitra lasiophthalma to 14.3% of seven Hybomitra epistates. Infected I. dammini larvae and nymphs coexisted with infected Dermacentor variabilis (American dog tick)immatures on white-footed mice (Peromyscus leucopus), but unlike I. dammini, none of the 55 adult American dog ticks collected from vegetation harbored B. burgdorferi. Groups of 113 field-collected mosquitoes ofAedes canadensis and 43 Aedes stimulans were placed in cages with uninfected Syrian hamsters. Of these, Il females of both species contained B. burgdorferi and had fed fully or partially from the hamsters. No spirochetes were isolated from the hamsters, but antibodies were produced in one test animal.

The causative agent of Lyme disease, Borrelia burgdorferi, has been detected in or isolated from ticks (2, 5, 9-11, 21), human tissues (8, 32), and the blood and organs of wildlife (1-3, 5, 19). Although Ixodes dammini is the chief vector of B. burgdorferi in the northeastern United States, Wisconsin, and Minnesota, other arthropods, such as mosquitoes, horse flies, and deer flies, have also been found harboring this bacterium (22). Evidence of transmission by hematophagous insects, however, is limited to infrequent associations between bites and the development of erythema migrans (13, 15, 22), a unique skin lesion that marks the earlystage of Lyme disease (31, 32).

Members of several species of arthropods harbor B. burgdorferi during the summer, but little is known about the prevalence of infection in different sites and years. Moreover, it is unclear whether naturally infected mosquitoes, horse flies, and deer flies retain spirochetes in their digestive tracts, complete gonotrophic cycles, and efficiently transmit B. burgdorferi to vertebrate hosts. The objectives of this study were to compare prevalences of infection for ticks and biting insects in two rural communities in Connecticut where human infections with Lyme disease have been documented, to determine reproductive life histories for spirochete-infected blood-seeking mosquitoes, horse flies, and deer flies,to determine whether B. burgdorferi can survive in experimentally infected horse flies, and to attempt to isolate B. burgdorferi from Syrian hamsters fed upon by naturally infected mosquitoes in the laboratory.

MATERIALS AND METHODSStudy sites and sampling. Ticks, mosquitoes, deer flies, and horse flies were collected during 1986 and 1987 in Salem and Norwich, communities in southeastern Connecticut where Lyme disease is endemic (23, 33). During spring and summer, immatures of I. dammini and Dermacentor variabilis (American dog tick) were removed from Peromyscus leucopus (white-footed mouse) captured in or near forests, while adult ticks of both species were obtained during spring or summer by flagging vegetation along trails near woodlands. Blood-seeking mosquitoes and deer flies were captured in an insect net during the summer (time of collection,1000 to 1500 h) as they approached the investigator. Mosquitoes were removed by an aspirator and separated from deer flies before being transported to the laboratory. Horse flies were collected during June and July by erecting dry-icebaited canopy traps (28). All insects were kept on crushed ice or in styrofoam containers in the field and while in transit.Dissection and identification of B. burgdorferi. Midgut tissues were dissected from ticks and smeared onto glass microscope slides as described previously (5, 9). Heads were removed from mosquitoes and placed on other microscope slides. Internal contents were expelled and smeared by applying pressure to cover slips placed over the heads.Anterior digestive tracts, including the salivary glands and proventriculus, were dissected from field-collected horse flies and deer flies (tabanids) and from experimentally infected females of a salt marsh horse fly, Tabanus nigrovittatus. These tissues were smeared onto slides in the same manner as the tick and mosquito tissues. After drying at 37°C, all preparations were fixed in acetone for 10 min and overlaid with murine monoclonal antibody (H5332) diluted 1: 4 or 1:8 in phosphate-buffered saline (PBS) solutions. This antiserum was directed to outer surface protein A, a polypeptide of approximately 31 kilodaltons (6, 7) that is common to all North American isolates of B. burgdorferi. The specificity of this monoclonal antibody, dilutions of reagents, application of fluorescein isothiocyanate-labeled goat anti-mouse immunoglobulin G (IgG) (1:40), and other procedures used in indirect fluorescent-antibody (IFA) staininghave been reported (5-7, 21, 22). Reproductive life histories. Ovaries were dissected from mosquitoes and tabanids to determine the number of completed gonotrophic cycles. Tissues were teased apart in Ringer physiological saline solution, and ovarioles were examined microscopically to determine whether dilatations or relics (yellow bodies) had formed in follicular tubes. The presence of these structures is evidence of egg development1482Vol. 26, No. 8Downloaded from jcm.asm.org by on November 3, 2009B. BURGDORFERI IN TICKS AND INSECTS 1483 and oviposition (parity) and can be used to determine the number of ovarian cycles completed. Dissection procedures, terminology for ovarian structures, and interpretations of parity are those of Detinova (12). In general, females of most biting insects ingest blood from vertebrate hosts, maturetheir eggs, and deposit them in or near aquatic or semiaquatic habitats. Multiple blood feedings and ovarian cycles can occur (12, 20, 34).Feeding trials and isolation attempts. Blood-seeking mosquitoes of Aedes canadensis and Aedes stimulans were collected from woodlands in Salem and Norwich. Groups of females, separated by species, were placed into screened cages with anesthetized, uninfected Syrian hamsters in thelaboratory. These mammals can be used to isolate B. burgdorferi (17). All test animals were initially immobilized with Penthrane (Abbott Laboratories, North Chicago, Ill.) and subsequently injected with ketamine hydrochloride (Vetalar; Parke-Davis, Morris Plains, N.J.). Hamsters were shaved along the head and back to allow mosquitoes to feed readily.After 1 h, the blood-engorged mosquitoes were separated from the unfed individuals, dissected, and screened for spirochetes by IFA staining methods. Two additional hamsters were each inoculated intraperitoneally with 150 ,ul of Barbour-Stoenner-Kelly (BSK) medium containing living B.burgdorferi (CT strain 22956), a 10-day-old culture of a primary isolate recovered from a kidney of P. leucopus captured in Armonk, N.Y. The number of spirochetes in the culture medium was about 4.0 x 106/ml. For negative controls, four normal hamsters (not exposed to mosquitoes) were kept in separate cages adjacent to those of the hamsters fed upon by mosquitoes or challenged by inoculation. Kidneys and spleens were aseptically removed, triturated, and inoculated into BSK medium as described previously (1, 3, 17). Cultures were held at 31°C for 4 to 7 weeks, and samples of medium were examined by dark-field microscopyto detect living spirochetes. In addition, blood samples were collected from hamsters, and sera were stored at -60°C until IFA analyses for antibodies to B. burgdorferi could be performed.Laboratory experiments were conducted to determine whether B. burgdorferi survives in females of T. nigrovittatus. Blood-seeking horse flies were collected in canopy traps during July 1985 and August 1986 in a salt marsh in Milford, Conn. After being transferred to sugar-free cages inthe laboratory, the insects were held overnight with distilled water at 21 + 3°C. On the next day, they were placed on membrane feeding devices (26) for 20 to 30 min. The food source was kept in a glass reservoir and consisted of 2.0 ml of fresh, citrated beef blood mixed with equal volumes of 7-day-old BSK medium containing living B. burgdorferi (CT strain 2591) or PBS (negative control). Following feeding trials, females were dissected at hourly or daily intervals, and duplicate preparations of head and digestive tract tissues were examined for spirochetes by dark-field microscopy and IFA staining methods.Serologic testing. Hamster serum samples were screened for total immunoglobulin to B. burgdorferi by IFA staining procedures (5, 21). Polyvalent fluorescein isothiocyanatelabeled goat anti-hamster IgG (Kirkegaard & Perry Laboratories, Gaithersburg, Md.) was diluted 1:30 in PBS solution. Sera from inoculated hamsters, obtained during earlier experiments, served as positive controls; B. burgdorferi infections were confirmed by reisolating the spirochetes from kidney or spleen tissues. Uninfected (normal) hamster sera were also included as controls. In preliminary analyses of nine normal serum samples, there was no nonspecific reac-TABLE 1. Total arthropods collected and number of specimens infected with B. burgdorferi in Salem and Norwich, Conn., during 1986 and 1987Salem NorwichArthropod species No. of Nfo. o of No specimens infected specimens infected collected collectedTicksbI. dammini 116 42 (36.2) 2 0D. variabilis 52 10 (19.2) 14 0MosquitoesA. canadensis 113 11 (9.7) 10 0A. cinereus 19 0 5 0A. stimulans 21 2 (9.5) 23 1 (4.4)A. triseriatus 18 2 (11.1) 36 0Deer fliesC. callidus 106 9 (8.5) 57 6 (10.5)C. cincticornis 20 0 9 0C. geminatus 49 0 0 CC. macquarti 36 3 (8.3) 2 0C. univittatus 144 13 (9.0) 1 0C. vittatus 101 3 (3.0) 10 0Horse fliesH. epistates 7 1 (14.3) 1 0H. hinei 51 2 (3.9) 17 0H. lasiophthalma 101 8 (7.9) 105 3 (2.9)H. sodalis 38 0 0T. lineola 15 0 0T. quinquevittatus 12 0 2 0a Determined by IFA staining with murine monoclonal antibody H5332.b Larvae and nymphs removed from white-footed mice.c-, Not determined.tivity at dilutions of -1:16. Therefore, reactions of test seraat or above a 1:16 dilution were considered positive. Allserum samples were retested to determine reproducibility.Statistical analyses. When sample sizes were adequate (n >30), variances were computed and tested for homogeneity byan F test (30). Statistical differences in sample means werethen determined by an appropriate Student's t test. Allanalyses were conducted at the P < 0.01 level of significance.RESULTSTicks, mosquitoes, and tabanids of 12 species harbored B.burgdorferi. The percentage of infected I. dammini immatures(36.2%) removed from white-footed mice in Salem wassignificantly greater than those of all other arthropods studied(Table 1). The second highest rate (19.2%) was recordedfor immatures of D. variabilis. Spirochetes in midgut preparationsof I. dammini usually exceeded an average of 50 per40x microscopic field. Tissues of naturally infected horseflies and deer flies normally contained ca. 10 to 50 spirochetesper field, while after considerable searching, preparationsof head tissues from mosquitoes rarely exceeded 15spirochetes per field. When present, B. burgdorferi wasfound most readily in tissues from I. dammini. In Norwich,females of A. stimulans, Chrysops callidus, and H. lasiophthalmaalso contained B. burgdorferi; the prevalence ofinfection was 10.5% or less.Examinations of ovarian tissues from 1,062 blood-seekingmosquitoes and tabanids revealed that the majority hadcompleted at least one gonotrophic cycle and, therefore, hadtaken at least one blood meal before depositing eggs (oviposition)and starting the second ovarian cycle. For uninfectedinsects, the numbers of parous specimens (i.e., with evi-VOL. 26, 1988

To determine whether mosquitoes could transmit B. burgdorferi,field-caught blood-seeking females of A. canadensisand A. stimulans were allowed to feed on uninfected hamstersin the laboratory. Five groups of 113 A. canadensisfemales were each placed with five separate hamsters (16 to35 mosquitoes per group), while two groups of 43 A. stimulansfemales (15 and 28 per group) were placed with twoother hamsters. Of these, 71 A. canadensis and 30 A.stimulans ingested partial or complete blood meals. B.burgdorferi was detected in the head tissues of nine and twoblood-fed specimens, respectively. At least one infectedmosquito had fed partially or completely from each of theexposed hamsters. No isolations were made from thespleens or kidneys of seven hamsters fed upon by mosquitoesor from four others held as negative controls. 'However,B. burgdorferi was recovered from the two hamsters thathad been inoculated with BSK medium containing thesespirochetes. In addition, one of five hamsters fed upon by A.canadensis had antibodies to B. burgdorferi at a titer of 1:32.An infected mosquito ingested blood from this hamster, andthe antibody titer was reproducible. The remaining hamstersexposed to mosquitoes or held as negative controls had noantibodies. Those inoculated with spirochetes had antibodytiters of 1:64 or 1:256.

DISCUSSIONMembers of several species of arthropods harbor B.burgdorferi, but the prevalence of infection was highlyvariable. In I. dammini, the chief vector of B. burgdorferi inConnecticut (4, 5, 21, 32), the proportion of infected ticksdiffered from 11 to 54%, depending on the site, season, andsampling method. At Shelter Island, N.Y., an infection rateof 61% has been reported (9). The presence of B. burgdorferiin biting insects also varied. During 1985, 14 species ofhematophagous insects were found to be carrying this bacteriumin Norwich, Conn. (22); rates were as high as 21% forC. callidus. In the present study, females of three speciesfrom Norwich contained B. burgdorferi, and the prevalenceof infection for C. callidus was 50% lower.

Horse flies and deer flies can disperse readily from breedingareas (16, 35). The infected females collected in Norwichmay have acquired B. burgdorferi elsewhere. This study sitedoes not appear to be an important focus for Lyme disease,because the numbers of I. dammini on mice were very lowand B. burgdorferi was not found in these ticks. In addition,we did not collect I. dammini adults while flagging vegetationduring the spring and fall. Although the sources ofinfection for mosquitoes and tabanids are unknown, thenumber of infected specimens may vary with changes inpopulation densitites of large mammals such as white-taileddeer (Odocoileus virginianus), horses, or cattle. Antibodiesto B. burgdorferi have been detected in deer (21, 24) andhorses (27; L. A. Magnarelli, J. F. Anderson, E. Shaw, J. E.Post, and F. C. Palka, Am. J. Vet. Res., in press), indicatingthat these animals were exposed to the Lyme disease spirocheteor to another closely related Borrelia organism. However,isolation and identification of B. burgdorferi areneeded to confirm that the large mammals are spirochetemicand serve as reservoirs of infection. In comparison, whitefootedmice are abundant in forests and known to becompetent reservoirs for B. burgdorferi (2, 3, 14, 18). Withrelatively small home ranges, and as important hosts for I.dammini, these rodents serve to maintain B. burgdorferiinfections in foci and to infect ticks during the warmermonths. Therefore, the presence of infected I. dammini isconvincing evidence that the sampling area is a focus forLyme disease.Infected I. dammini and D. variabilis coexisted on whitefootedmice. This reinforces the epidemiological significanceof this rodent in Lyme borreliosis. Since transovarial transmissionof B. burgdorferi is low in I. dammini (25, 29), larvaemainly acquire these spirochetes by feeding on infectedhosts. Based on lower percentages of infected D. variabilisnymphs and the absence of B. burgdorferi in questing adultsof this species, transstadial transmission in D. variabilis isprobably inefficient. In addition, there are no convincingreports indicating an association between American dog tickbites and the development of erythema migrans in humans.Therefore, adults of this species do not appear to be vectorsof B. burgdorferi.The occurrence of infected ticks and biting insects inSalem indicates that B. burgdorferi is widely distributedamong hematophagous arthropod populations. As in EastHaddam and Lyme, two communities that border Salemwhere Lyme disease is also endemic (23, 33), I. dammini isabundant. Verification of B. burgdorferi in arthropods fromSalem confirms earlier clinical reports on human infectionsin this community. Since birds carry infected I. damminilarvae and nymphs (4), the range of this tick may continue toexpand, and Lyme disease may become endemic in othercommunities in southcentral and southeastern Connecticut.

Ovarian examinations of biting insects revealed thatnearly all of the infected, blood-seeking mosquitoes andtabanids had completed at least one ovarian cycle. Unliketicks, mosquitoes and tabanids can ingest multiple bloodmeals from different mammals during a gonotrophic cycle(20, 34) and in the process may acquire B. burgdorferi fromone or more infected hosts. Maximal periods of survival forB. burgdorferi in naturally infected biting insects are unknown,but in experimentally infected mosquitoes, B. burgdorferilived less than 6 days in the insect's digestive system(26). This, coupled with the relatively low number of spirochetesfound in head tissues of field-caught females and noisolates of B. burgdorferi from hamsters fed upon by infectedmosquitoes, indicates that these insects may not be suitablehosts for this bacterium. The low-level immune response ina hamster fed upon by an infected female of A. canadensismay have been directed against dead or weakened B. burgdorferi.Although anecdotal, there are records of deer flybites and the subsequent development of erythema migransin persons who had Lyme disease (22). In addition, B.burgdorferi survives for brief periods in T. nigrovittatus.Further studies are needed to confirm that deer flies or horseflies can mechanically transmit B. burgdorferi.

Abstract
Millions of Lyme disease vector ticks are dispersed annually by songbirds across Canada, but often overlooked as the source of infection. For clarity on vector distribution, we sampled 481 ticks (12 species and 3 undetermined ticks) from 211 songbirds (42 species/subspecies) nationwide. Using PCR, 52 (29.5%) of 176 Ixodes ticks tested were positive for the Lyme disease spirochete, Borrelia burgdorferi s.l. Immature blacklegged ticks, Ixodes scapularis, collected from infested songbirds had a B. burgdorferi infection prevalence of 36% (larvae, 48%; nymphs, 31%). Notably, Ixodes affinis is reported in Canada for the first time and, similarly, Ixodes auritulus for the initial time in the Yukon. Firsts for bird-parasitizing ticks include I. scapularis in Quebec and Saskatchewan. We provide the first records of 3 tick species cofeeding on passerines (song sparrow, Swainson's thrush). New host records reveal I. scapularis on the blackpoll warbler and Nashville warbler. W e furnish the following first Canadian reports of B. burgdorferi-positive ticks: I. scapularis on chipping sparrow, house wren, indigo bunting; I. auritulus on Bewick's wren; and I. spinipalpis on a Bewick's wren and song sparrow. First records of B. burgdorferi-infected ticks on songbirds include: the rabbit-associated tick, Ixodes dentatus, in western Canada; I. scapularis in Quebec, Saskatchewan, northern New Brunswick, northern Ontario; and Ixodes spinipalpis (collected in British Columbia). The presence of B. burgdorferi in Ixodes larvae suggests reservoir competency in 9 passerines (Bewick's wren, common yellowthroat, dark-eyed junco, Oregon junco, red-winged blackbird, song sparrow, Swainson's thrush, swamp sparrow, and white-throated sparrow). We report transstadial transmission (larva to nymph) of B. burgdorferi in I. auritulus. Data suggest a possible 4-tick, i.e., I. angustus, I. auritulus, I. pacificus, and I. spinipalpis enzootic cycle of B. burgdorferi on Vanco uver Island, British Columbia.

Our results suggest that songbirds infested with B. burgdorferi-infected ticks have the potential to start new tick populations endemic for Lyme disease. Because songbirds disperse B. burgdorferi-infected ticks outside their anticipated range, health-care providers are advised that people can contract Lyme disease locally without any history of travel.

Anaplasma phagocytophilum, the causative agent of human granulocytic anaplasmosis, is an obligate intracellular bacterium most commonly acquired from tick bites. High seroprevalence rates in endemic regions suggest that transfusion transmission of A phagocytophilum would be a common event; however, only 2 cases have previously been reported. The exact cause of this discrepancy is not known. Whole blood leukocyte-reduction methods used by many blood centers are thought to reduce the risk of transfusion transmission of many pathogens, including A phagocytophilum. We report 2 additional cases of transfusion-transmitted A phagocytophilum in which leukocyte reduction of all transfused units failed to prevent microbial transmission.

Hennepin County Medical Center and the University of Minnesota Medical School, Minneapolis, Minnesota; Yale University School of Medicine, NewHaven, Connecticut.

From the text :"We report the case of a woman who developed Lyme discase during the first trimester of pregnancy. She did not recive antibiotic therapy. Her infant, born at 35 weeks gestational age, died of congenital heart disease during the first week of life. Histologie examination of autopsy material showed the Lyme disease spirochete in the spleen, kidneys, and bone marrow."

"A 28-year-old mother of two healthy children became pregnant for the third time in September 1983. Soon thereafter, she participated in outdooractivities in an area of northwestern Wisconsin known to be endemic for Lyme disease (3).

On 7 November 1983, she noted an expanding annular skin lesion in the left poplileal region reaching a size of 20 X 30 cm. She also developed two secondary skin lesions, headache, stiff neck, arthralgias, malaise, and inguinal lymphadenopathy. All symptoms resolved within severalweeks without treatment. Thereafter, the antepartum course was normal except for recurrent arthralgias during the third trimester. No medications were taken during the pregnancy.

On 6 May 1984, the patient delivered a 3000-g male infant whose estimated gestational age was 35 weeks. No skin lesions were seen. The baby had respiratory distress. An echocardiogram and cardiac catheterization showed a dilated. poorly contractile left ventricle; aortic valvular stenosis; patent ductus arteriosus; and coarctation of thc aorta. Despite emergency balloon catheter dilatation of the coarctation and aortic valvotomy, the infant died after 39 hours."

Location: From the Blood Bank and Transfusion Medicine, the Division of Infectious Disease, The Miriam Hospital, and the Rhode Island Blood Center, Providence, Rhode Island; and Imugen, Inc., Norwood, Massachusetts.

DOI: 10.1111/j.1537-2995.2012.03685.x

BACKGROUND: Human granulocytic anaplasmosis (HGA) is a tick-borne rickettsial infectious disease. To date four cases of transfusion-transmitted anaplasmosis (TTA) have been described in the literature, and only one from leukoreduced red blood cells (RBCs).

CASE REPORT: A 64-year-old patient with acute gastrointestinal blood loss was admitted to the hospital and received 5 units of prestorage leukoreduced RBCs. He was stabilized and discharged. He developed headache, fever, and chills 2 days after discharge and was readmitted. On Day 5 of his second admission polymorphonuclear leukocytes containing morulae consistent with HGA were reported in the peripheral smear.

RESULTS: Samples from the recipient tested positive by polymerase chain reaction (PCR) for Anaplasma phagocytophilum, the causative agent of HGA and a segment from one of the five donors tested positive by both serology and PCR.

Novel Animal Reservoir for Group of Tick-Borne Diseases Discovered -- And It Lives in Your Backyard

ScienceDaily (June 23, 2012) ? A team of scientists at Washington University in St. Louis has been keeping a wary eye on emerging tick-borne diseases in Missouri for the past dozen years, and they have just nailed down another part of the story.
See Also:
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Immune System

Plants & Animals

Spiders and Ticks
New Species

Earth & Climate

Exotic Species
Rainforests

Reference

Vector (biology)
Tick
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They knew from earlier work that the animal reservoirs for the diseases included white-tailed deer, wild turkey and a species in the squirrel famiiy, but the DNA assay they had used wasn't sensitive enough to identify the species.

Squirrels belong to a large family called the Sciuridae, which includes chipmunks, fox squirrels, red squirrels, flying squirrels, ground hogs and prairie dogs.

In the May issue of the Journal of Medical Entomology the scientists, led by Robert E. Thach, PhD, professor of biology in Arts & Sciences, report that a more sensitive assay has allowed them to identify the major species in question as the eastern gray squirrel.

Yes, the friendly neighborhood seed thief and dog tease is also a mobile tick blood supply and bacteria incubator.

The work is important because tick-borne diseases can be efficiently controlled only if all of the animal reservoirs that might contribute to transmission of the disease have been identified.

Not your New England tick

The most prevalent tick-borne disease in North America is Lyme disease, which is transmitted by the bite of an infected black-legged tick. In the southeastern United States, however, the most common diseases are ehrlichioses and STARI, which are transmitted by the bite of a different tick, the lone star tick.

Until 1986, ehrlichia bacteria were thought to cause disease only in animals. But in that year, a physician noticed mulberry-shaped aggregates characteristic of the bacteria in the blood of a gravely ill man.

The lone star tick, similarly, was thought to be merely a nuisance species until 1993, when the DNA of one of the ehrlichia species was found in lone star ticks collected in Missouri and several other states.

Ehrlichiosis typically begins with vague symptoms that mimic those of other bacterial illnesses. In a few patients, however, it progresses rapidly to affect the liver and other organs, and may cause death unless treated with antibiotics. STARI is similar to Lyme disease but seems to be less virulent.

The reservoirs

By 2010, with the pathogens and their vector identified, the WUSTL team was trying to find the animal reservoirs.

Looking for pathogens and host species, they ran two assays on the ground-up ticks: one to identify the DNA of pathogens and the other to identify the DNA of animals that had provided blood meals.

The blood meal assay on ticks carrying pathogens identified white-tailed deer blood and the blood of a species in the squirrel family, but it couldn't distinguish among 20 or so possible squirrel species.

So the team was very interested when they read a paper in the Journal of Medical Entomology about a new assay that could identify tick blood meals down to the species level.

The assay, developed by scientists at the University of Neuchatel in Neuchatel, Switzerland, used a segment of mitochondrial DNA instead of nuclear DNA as a species marker.

Mitochondria, organelles within the cells that convert energy into forms cells can use, have their own DNA, probably because they were once free-living bacteria.

For reasons that are not entirely clear, mitochondrial DNA mutates faster than DNA tucked away in the cell nucleus. It may be that the mitochondria simply have more primitive DNA repair mechanisms and so cannot fix mistakes if they occur.

In any case, the more mutations, the greater the difference between the DNA of two different species, and the greater the power of the assay to distinguish among species, Thach says.

To tailor the assay for their purposes, the team retrieved the DNA sequences for possible North American host species from Genbank, an open-access sequence database. Sequences not available in the database were determined by the lab.

Lisa S. Goessling, now a research lab supervisor in the School of Medicine, used the sequences to make a palette of probes for 11 species and -- just to make sure the net was cast wide enough -- several higher taxonomic orders.

The scientist then re-ran old samples and newly collected ticks through the new assay. Spots on the assay where the tick blood and the gray squirrel probe overlapped lit up, signaling the presence of gray squirrel blood in the ticks.

Why not the others?

Lone star ticks are famously aggressive and indiscriminate biters, so why hadn't they attacked other animals? Is there something special about deer or gray squirrels that makes the ticks prefer them?

This isn't the kind of question the scientists can answer definitively, but Thach doesn't think so. He has a simpler answer.

"If you think of an inventory of the animals in the woods and the amount of blood in each, well, most of the available blood in the woods is in deer, and next to that in turkeys and squirrels, because turkeys are so big and there are so many squirrels. So I suspect it's mainly just a mass phenomenon," he says.

Neighborhood, neighborhood, neighborhood

Having found gray squirrel DNA in tick blood, the scientists attacked the problem from a different angle to see if they could confirm their results. They trapped gray squirrels rather than ticks.

Were the gray squirrels carrying tick-borne pathogens? The answer, it turned out, depends on where you are. Only 5 percent of the squirrels in a relatively urban suburb (University City, Mo.) were carrying a pathogen, but 25 percent of the squirrels in a wooded "garden suburb" (Kirkwood, Mo.) were infected.

Why the difference? Thach suspects it comes down to white-tailed deer. There are few, if any, in University City, but they cruise backyards in Kirkwood. Wherever deer go they shed ticks.

This also is the likely answer to another conundrum: the absence or near absence of]ticks in Forest Park, the 1,371-acre urban park that adjoins Washington University in St. Louis. Thach says an exhaustive search turned up only one tick.

Why so few ticks? Perhaps because the only deer in Forest Park are the ones in the Saint Louis Zoo.

A 36-year-old woman acquired severe human granulocytic anaplasmosis after blood transfusion following a cesarean section. Although intensive treatment with mechanical ventilation was needed, the patient had an excellent recovery. Disease caused by Anaplasma phagocytophilum infection was confirmed in 1 blood donor and in the transfusion recipient.

Apr. 4, 2013 — Like humans, mice can become infected with Borrelia. However, not all mice that come into contact with these bacteria contract the dreaded Lyme disease: Animals with a particular gene variant are immune to the bacteria, as scientists from the universities of Zurich and Lund demonstrate. Wild mice are the primary hosts for Borrelia, which are transmitted by ticks.

Springtime spells tick-time. Lyme borreliosis is the most common tick-borne disease in Switzerland: around 10,000 people a year become infected with the pathogen. The actual hosts for Borrelia, however, are wild mice. Like in humans, the pathogen is also transmitted by ticks in mice. Interestingly, not all mice are equally susceptible to the bacterium and individual animals are immune to the pathogen. Scientists from the universities of Zurich and Lund headed by evolutionary biologist Barbara Tschirren reveal that the difference in vulnerability among the animals is genetic in origin.

Protective gene variant

Tschirren and colleagues examined wild mice for signs of a Borrelia infection in a large-scale field study. Borrelia afzelii -- the scientific name for the bacteria -- feed on mouse blood. The researchers discovered that mice with a particular variant of the antigen receptor TLR2 were around three times less susceptible toBorrelia. "The immune system of mice with this receptor variant recognizes the pathogen better and can trigger an immune response more quickly to destroy the Borrelia in time," says Tschirren. Infected mice exhibit similar symptoms to humans -- especially joint complaints. Consequently, in the wild infected mice probably do not survive for very long and weakened animals soon fall victim to foxes and birds of prey.

Arms race between mice and Borrelia

The protective gene variant is advantageous for its carriers and, according to the researchers, gradually becoming prevalent in the mouse population. Nonetheless, it is unlikely that all mice will one day be resistant to Borrelia. "The increasing resistance in the host is bound to lead to adaptations in Borrelia," predicts Tschirren. "We can observe the evolutionary adaptation through the rearmament in mice and the pathogen."

People also have the antigen receptor TLR2, but not the resistant gene variant observed in mice. Whether the evolutionary arms race between mice and Borrelia will have repercussions for people remains to be seen. According to Tschirren, the bacterium does not necessarily have to become more aggressive for humans.

The discovery of the key role of Toll-like receptors (TLRs) in initiating innate immune responses and modulating adaptive immunity has revolutionized our understanding of vertebrate defence against pathogens. Yet, despite their central role in pathogen recognition and defence initiation, there is little information on how variation in TLRs influences disease susceptibility in natural populations. Here, we assessed the extent of naturally occurring polymorphisms at TLR2 in wild bank voles (Myodes glareolus) and tested for associations between TLR2 variants and infection with Borrelia afzelii, a common tick-transmitted pathogen in rodents and one of the causative agents of human Lyme disease. Bank voles in our population had 15 different TLR2 haplotypes (10 different haplotypes at the amino acid level), which grouped in three well-separated clusters. In a large-scale capture–mark–recapture study, we show that voles carrying TLR2 haplotypes of one particular cluster (TLR2c2) were almost three times less likely to be Borrelia infected than animals carrying other haplotypes. Moreover, neutrality tests suggested that TLR2 has been under positive selection. This is, to our knowledge, the first demonstration of an association between TLR polymorphism and parasitism in wildlife, and a striking example that genetic variation at innate immune receptors can have a large impact on host resistance.FULL TEXT: http://rspb.royalsocietypublishing.org/ ... 30364.full

Transovarial transmission (TOT) of Borrelia burgdorferi (sensu lato), the agent of Lyme disease, by the Ixodes persulcatus group of hard ticks (Ixodidae) has frequently been reported in the literature since the discovery of Lyme disease 1982. Evidence for and against TOT by B. burgdorferi has led to uncertainty and confusion in the literature, causing misconceptions that may have public health consequences. In this report, we review the published information implicating B. burgdorferi as a bacterium transovarially transmitted among ticks of the Ixodes persulcatus group and present new data indicating the transovarially transmitted agent is actually Borrelia miyamotoi. B. miyamotoi, first described in 1995, is antigenically and phylogenetically related to B. burgdorferi, although more closely related to the relapsing fever-group Borrelia typically transmitted by soft ticks (Argasidae). Borrelia infections of unfed larvae derived from egg clutches of wild-caught Ixodes scapularis are demonstrated to result from transovarial transmission of B. miyamotoi, not B. burgdorferi. The presence of this second Borrelia species, apparently sympatric with B. burgdorferi worldwide also may explain other confusing observations reported on Borrelia/Ixodes relationships.

NEWS: Recent study suggests that Lyme disease can be sexually transmitted25th January 2014

spirochete image

Notes one researcher: "There is always some risk of getting Lyme disease from a tick bite in the woods. But there may be a bigger risk of getting Lyme disease in the bedroom.”

Press release, January 25, 2014:

Carmel, CA – A new study suggests that Lyme disease may be sexually transmitted. The study was presented at the annual Western Regional Meeting of the American Federation for Medical Research, and an abstract of the research was published in the January issue of the Journal of Investigative Medicine.

Lyme disease is a tick-borne infection caused by Borrelia burgdorferi, a type of corkscrew- shaped bacteria known as a spirochete (pronounced spiro’keet). The Lyme spirochete resembles the agent of syphilis, long recognized as the epitome of sexually transmitted diseases. Last summer the Centers for Disease Control and Prevention (CDC) announced that Lyme disease is much more common than previously thought, with over 300,000 new cases diagnosed each year in the United States. That makes Lyme disease almost twice as common as breast cancer and six times more common than HIV/AIDS.

“Our findings will change the way Lyme disease is viewed by doctors and patients,” said Marianne Middelveen, lead author of the study presented in Carmel. “It explains why the disease is more common than one would think if only ticks were involved in transmission.”

The present study was a collaborative effort by an international team of scientists. In addition to Middelveen, a veterinary microbiologist from Canada, researchers included molecular biologists Jennie Burke, Augustin Franco and Yean Wang and dermatologist Peter Mayne from Australia working with molecular biologists Eva Sapi and Cheryl Bandoski, family practitioner Hilary Schlinger and internist Raphael Stricker from the United States.

In the study, researchers tested semen samples and vaginal secretions from three groups of patients: control subjects without evidence of Lyme disease, random subjects who tested positive for Lyme disease, and married heterosexual couples engaging in unprotected sex who tested positive for the disease.

As expected, all of the control subjects tested negative for Borrelia burgdorferi in semen samples or vaginal secretions. In contrast, all women with Lyme disease tested positive for Borrelia burgdorferi in vaginal secretions, while about half of the men with Lyme disease tested positive for the Lyme spirochete in semen samples. Furthermore, one of the heterosexual couples with Lyme disease showed identical strains of the Lyme spirochete in their genital secretions.

“The presence of the Lyme spirochete in genital secretions and identical strains in married couples strongly suggests that sexual transmission of the disease occurs,” said Dr. Mayne.

“We don’t yet understand why women with Lyme disease have consistently positive vaginal secretions, whilst semen samples are more variable. Obviously there is more work to be done here.”

Dr. Stricker pointed to the unknown risks of contracting Lyme disease raised by the study. “There is always some risk of getting Lyme disease from a tickbite in the woods,” he said. “But there may be a bigger risk of getting Lyme disease in the bedroom.”

Reference: The Journal of Investigative Medicine 2014;62:280-281.

\Presented at the Western Regional Meeting of the American Federation for Medical Research, Carmel, CA, January 25, 2014. http://afmr.org/Western/.